Arthropod venoms: a vast arsenal of insecticidal neuropeptides

ReviewArthropod Venoms: A Vast Arsenal of Insecticidal Neuropeptides

Elisabeth F. Schwartz, Caroline B. F. Mourao, Karla G. Moreira, Thalita S. Camargos,Marcia R. MortariDepartamento de Ciencias Fisiologicas, Laboratorio de Toxinologia, Universidade de Brasılia, Brasılia, DF 70910-900, Brazil

Received 20 January 2012; revised 9 May 2012; accepted 23 May 2012

Published online 7 June 2012 in Wiley Online Library (wileyonlinelibrary.com). DOI 10.1002/bip.22100

This article was originally published online as an accepted

preprint. The ‘‘Published Online’’ date corresponds to the

preprint version. You can request a copy of the preprint by

emailing the Biopolymers editorial office at biopolymers@wiley.

com

INTRODUCTION

Highlighting the Presence of Components with

Insecticidal Activity from the Venom of Arthropods

The phylum Arthropoda is the most abundant and

diverse, totaling about 75% of the world’s animal

species. Arthropods have a wide distribution, occupy-

ing virtually every ecological niche on the planet.

Over millions of years of evolution, venomous

arthropods have developed an arsenal of bioactive com-

pounds that paralyzes and/or kills other organisms, with an

important role in capturing food and defending against

predators. Arthropod venoms are formed by a variety of bio-

logically active compounds, such as proteins and peptides,

acylpolyamine toxins, and biogenic amines.1–4

Currently, the venoms of arthropods have been recognized

as one of the greatest resources of biologically active mole-

cules, although most of this arsenal remains unknown.5 Acyl-

polyamines and peptides are the main representatives of neu-

rotoxins and because of their action; they have been used as

tools to extend the knowledge of molecular neurotransmis-

sion mechanisms.2,6–8 The study of venom neurotoxins has

also led to the discovery of novel natural insecticides, as well

as the development of tools for the study of drugs potentially

useful for the treatment of neurological disorders.7,9–11

Although many molecular targets have not been satisfac-

torily investigated yet, neuroactive peptides isolated from

these animals can act on ion channels, membrane receptors,

and neurotransmitter transporters, with high affinity and

specificity toward the nervous system of invertebrates. The

great difficulty in identifying the molecular targets of

ReviewArthropod Venoms: A Vast Arsenal of Insecticidal Neuropeptides

Correspondence to: Marcia Renata Mortari, Laboratorio de Toxinologia, Departa-

mento de Ciencias Fisiologicas, Instituto de Ciencias Biologicas, Universidade de

Brasılia, Brasılia, 70910-900, DF, Brasil; e-mail: [email protected]

ABSTRACT:

Arthropods are the most diverse animal group on the

planet, and occupy almost all ecological niches. Venomous

arthropods are a rich source of bioactive compounds

evolved for prey capture and defense against predators

and/or microorganisms. These highly potent chemical

arsenals represent an available source for new insecticidal

compounds as they act selectively on their molecular

targets. These toxins affect the invertebrate nervous

system and, until the moment, several insecticidal

compounds belonging to the class of peptides or

polyamine-like compounds have been purified and

characterized from the venom of arachnids and

hymenopterans. This review focuses on invertebrate-

specific peptide neurotoxins that have been isolated from

the venom of spiders, scorpions, centipedes, ants, and

wasps, discussing their potential in pest control and as

invaluable tools in neuropharmacology. # 2012 Wiley

Periodicals, Inc. Biopolymers (Pept Sci) 98: 385–405,

2012.

Keywords: insecticidal peptides; arthropod venom; spider;

scorpion; hymenopteran

Contract grant sponsor: CNPq

Contract grant number: 303003/2009-0 and 564223/2010-7

Contract grant sponsor: FAPDF

Contract grant number: 193.000.472/2008

VVC 2012 Wiley Periodicals, Inc.

PeptideScience Volume 98 / Number 4 385

peptides may be mainly due to the limited amount of venom,

as well as the absence of several subtypes of ion channels and

biological models for experimentation, and also, in some

cases, the lack of specific agonists and antagonists to discrim-

inate the different subtypes of ion channels.

This review deals with insecticidal peptides from arthropod

venoms: arachnids (spiders and scorpions), Hymenoptera

(wasps and ants), and centipedes. All these animals use their

venom to catch prey, defend themselves against predators, and

some may also use for intraspecific communication. Table I

shows most of insect selective toxins with low or no toxicity to

mammals isolated from these animal venoms known to date.

Spiders belong to the order Araneae (Arachnida) and are

one of the most diverse groups of terrestrial animals.3 Several

neurotoxins of spider venom have been identified,12 charac-

terized mainly by peptides and acylpolyamines, and these

classes of molecules represent about two thirds of the dry

weight of spider venoms.13 So far, there is about 100 spiders’

peptide toxins described on the ArachnoServer database14

with action on insects of different orders (Lepidoptera, Dip-

tera, Orthoptera, Blattaria and Coleoptera). While most of

them were tested against insects belonging to only one order,

16 were toxic to insects of two or more orders, and some are

also toxic to mammals. The molecular target of most of these

insecticidal toxins are the calcium or sodium voltage-gated

channels (Cav and Nav, respectively) or, to a lesser extent, the

potassium channels activated by calcium or the ionotropic

glutamate receptors. However, *50% of these toxins do not

have their molecular targets characterized yet.14

Scorpions were the first arthropods to conquer the terres-

trial environment, more than 400 million years ago. They are

animals that survive in extreme and adverse physical condi-

tions, with nocturnal habits and remain hidden during the

day, feeding on small insects. Scorpion venom is a rich source

of different compounds such as enzymes, peptides, nucleo-

tides, lipids, and unknown compounds. The best-known tox-

ins are those specific for Na+ or K+ channels (NaScTxs and

KTxs, respectively).15,16 These peptides act on ion channels

with high affinity and specificity, and may also discriminate

between ion channels of insects and mammals.

Spider and scorpion insecticidal neurotoxins are up to 9

kDa peptides with a high content of disulfide bonds, which

increase their in vivo stability and resistance to proteases,

leading to a decrease in the degradation rates in the venom

gland and in the prey. These peptides cause death or rapidly

paralyze the prey by interacting with ion channels and/or

receptors in the neuromuscular system.17–19 Due to the stable

structure of these molecules, they can be quickly transported

via diffusion in the hemolymph of insects, reaching their

molecular target.20

Hymenoptera (including bees, wasps and ants) are ex-

traordinarily prevalent and widespread over the Earth and

they have been survived for at least 300 million years.21 The

great majority of species of both social and solitary wasps

are predators, including many agricultural pests. In general,

the venom of Hymenoptera is composed mainly of proteins,

peptides, biogenic amines, and inorganic salts.22 Among the

high molecular weight compounds there are many types of

allergens, such as antigens 5,23–25 hyaluronidases,26 and

phospholipases A1 and A2.27–29 In addition, recent studies

report that *70% of venom of social wasps consists of pep-

tides,30,31 which have been classified into polycationic pep-

tides encompassing the mastoparans, kinins, and chemotac-

tic peptides.22,32,33 In solitary wasp venom, the main com-

ponents are acylpolyamines, proteins and, similarly to

social wasps, neuroactive peptides. Conversely, it is impor-

tant to note the presence of melittin, apamin, adolapin, ter-

tiapin, and mast cell degranulating peptides in bee

venom.34,35

Most of predatory ants capture their prey using their ven-

oms. Like wasps, ant venoms consist of proteins, peptides

and a mixture of low-molecular mass compounds. Two ant

peptides are so far described with insecticidal activity: poner-

icins36 and poneratoxins.37 Ponericins, isolated from the

venom of the predatory ant Pachycondyla goeldii, exhibit in-

secticidal activity against cricket.36

Centipedes are among the earliest terrestrial animals and

their venoms have been poorly characterized so far. Ester-

ases, proteinases, alkaline and acid phosphatases, cardiotox-

ins, histamine, and neurotransmitter releasing compounds

were reported in Scolopendra venoms.38–40 A Scolopendra

sp. venom fraction (SC1) induced an increase in the leak

current on the cockroach giant axon, which was correlated

with the decrease in the membrane resistance.41 Rates

et al.42 observed that the venom of Scolopendra viridicornis

nigra causes legs and wings retraction in houseflies and is

capable of instantly paralyzing cockroaches (Periplaneta

americana) and crickets (Acheta domesticus). Until now,

there are no reports of insecticidal peptides isolated from

the Scolopendra genus venom.

The arthropod insecticidal peptides described to date are

presented here according to their targets and mechanisms of

action. We briefly point out their structural diversity and

their potential use in the agribusiness. Due to species selec-

tivity, in the last few decades, it has grown the identification

of insect-selective toxins that can be used to develop biologi-

cal recombinant pesticides as a safer substitute to broad-

spectrum chemical insecticides.17,18,43,44

386 Schwartz et al.

Biopolymers (Peptide Science)

TableI

ArthropodPeptides

withInsecticidalActivity

Arthropod

Peptide

Source

Target

Assay

Dose

UNIPROT

Spider

HNTX-I(l-TRTX-H

hn2b)

Haplopelmahainanum

InsectNa v

channel

IC50

4.36

0.3lM

(para/tipE)68

D2Y1X6

Tx4(6–1)(d-C

NTX-Pn1a)

Phoneutrianigriventer

InsectNa v

channel

LD50

3.86

2ng/house

fly(M

uscadom

estica)69

P59368

Magi-2(l-H

XTX-M

g1a)

Macrothelegigas

InsectNa v

channel

LD50

17.6nmolg�

1(Spodoptera

litura)55

P83558

Magi-3(l-H

XTX-M

g2a)

Macrothelegigas

InsectNa v

channel

LD50

>32.8nmolg�

1(S.litura)55

P83559

d-PaluIT1(d-A

MATX-PI1a)

Pireneitega

luctuosa

InsectNa v

channel

LD50

9.56

3.7lg

g�1(S.litura)81

P83256

d-PaluIT2(d-A

MATX-PI1b)

Pireneitega

luctuosa

InsectNa v

channel

LD50

24.76

11.18lg

g�1(S.litura)81,82

P83257

d-PaluIT3(d-A

MATX-PI1c)

Pireneitega

luctuosa

InsectNa v

channel

LD50

12.36

5.0lg

g�1(S.litura)81

P83258

d-PaluIT4(d-A

MATX-PI1d)

Pireneitega

luctuosa

InsectNa v

channel

LD50

>44.8lgg�

1(S.litura)81

P83259

l-Aga-I(l-AGTX-A

a1a)

Agelenopsisaperta

InsectNa v

channel

LD50

286

7lg

g�1(M

anduca

sexta)84

P11057

l-Aga-II(l-AGTX-A

a1b)

Agelenopsisaperta

InsectNa v

channel

LD50

756

27lgg�

1(M

.sexta)84

P11058

l-Aga-III(l-AGTX-A

a1c)

Agelenopsisaperta

InsectNa v

channel

LD50

286

12lgg�

1(M

.sexta)84

P60178

l-Aga-IV(l-AGTX-A

a1d)

Agelenopsisaperta

InsectNa v

channel

LD50

406

9lgg�

1(M

.sexta)84

P11060

l-Aga-V

(l-AGTX-A

a1e)

Agelenopsisaperta

InsectNa v

channel

LD50

486

11lgg�

1(M

.sexta)84

P11061

l-Aga-V

I(l-AGTX-A

a1f)

Agelenopsisaperta

InsectNa v

channel

LD50

386

12lgg�

1(M

.sexta)84

P11062

curtatoxinI(l-AGTX-H

c1a)

Hololenacurta

InsectNa v

channel

LD50

20lgg�

1(Achetadom

estica)86

P15967

curtatoxinII(l-AGTX-H

c1b)

Hololenacurta

InsectNa v

channel

LD50

4lg

g�1(A

.dom

estica)9,86

P60177

curtatoxinIII(l-AGTX-H

c1c)

Hololenacurta

InsectNa v

channel

LD50

4lgg�

1(A

.dom

estica)86

P15968

J-ACTX-H

v1a(j-H

XTX-H

v1a)

Hadronycheversuta

InsectBKCa

LD50

3036

42pmolg�

1(A

.dom

estica)129

P82227

J-ACTX-H

v1b(j-H

XTX-H

v1b)

Hadronycheversuta

InsectBKCa

LD50

2146

16pmolg�

1(A

.dom

estica)129

P82226

J-ACTX-H

v1c(j-H

XTX-H

v1c)

Hadronycheversuta

InsectBKCa

LD50

1676

10pmolg�

1(A

.dom

estica)128,129

P82228

j-TRTX-Ec2a

Eurocratoscelusconstrictus

InsectBKCa

IC50[peak

I BK(C

a)]

3.7nM

(Gryllusbimaculatus)133

–

j-TRTX-Ec2b

Eurocratoscelusconstrictus

InsectBKCa

IC50[peak

I BK(C

a)]

25.3nM

(G.bimaculatus)133

–

x-ACTX-H

v1a(x

-HXTX-H

v1a)

Hadronycheversuta

M-LVAandHVACa v

channels

LD50

0.38lg

g�1(A

.dom

estica),176,274

P56207

LD50

0.326

0.02lg

g�1(M

.dom

estica)274

IC50

279nM

(M-LVA)177

IC50

1080nM

(HVA)177

x-ACTX-A

r1a(x

-HXTX-A

r1a)

Atraxrobutus

M-LVAandHVACa v

channels

LD50

2366

28pmolg�

1(A

.dom

estica)177

P83580

IC50

692nM

(M-LVA)177

IC50

644nM

(HVA)177

x-ACTX-H

v2a(x

-HXTX-H

v2a)

Hadronycheversuta

HVACa v

channels

PD50

1606

9pmolg�

1(A

.dom

estica)178

P82852

EC50

*139pM

(bee

brain

neurons)178

HWTX-V

(x-TRTX-H

h2a)

Haplopelmaschmidti

HVACa v

channels

ED50

166

5lg

g�1(M

igratory

manieusis)179,180

P61104

IC50

2196

4nM

(cockroachDUM

neurons)180

PnTx4–3(d-C

NTX-Pn1b)


Glutamateneurotransm

ission

LD50

20ng/house

fly(M

.dom

estica)213

P84034

PnTx4(5–5)(c-C

NTX-Pn1a)


NMDAreceptor

LD50

9.3ng/house

fly(M

.dom

estica)275

P59367

Ba1

(U1-TRTX-Ba1a)

Brachypelmaruhnaui

ND

LD50

10.86

1.4lg

g�1(A

.dom

estica)240

P85497

Ba2

(U1-TRTX-Ba1b)

Brachypelmaruhnaui

ND

LD50

9.26

0.9lg

g�1(A

.dom

estica)240

P85504

Arthropod Venoms 387


Scorpion

a-anti-insectBjaIT

Buthotusjudaicus

Na v:site3

PD50

0.13lg

g�1(Sarcophaga

falculata)103

Q56TT9

aanti-insectLqhaIT

Leiurusquinquestriatushebraeus

Na v:site3

LD50

0.14lgg�

1(S.falculata)276

P17728

aanti-insectBotIT1

Buthusoccitanustunetanus

Na v:site3

LD50

0.6lg

g�1(Blatellagerm

anica)102

P55902

b-insectexcitatory

AaH

IT1

Androctonusaustralis

Na v:site4

LD50

7.6lg

g�1(S.litura)277

P01497

Kd

1–3nM

278

b-insectexcitatory

BmKIT1

Mesobuthusmartensii

Na v:site4

PD50

0.18lg

g�1(G

.bimaculatus)279

O61668

BotIT2


Na v:site4

LD50

3.5lgg�

1(B.germ

anica)280

P59863

Kd

0.36

0.1nM

281

b-depressantBotIT4


Na v:site4

LD50

1.1lgg�

1(B.germ

anica)102

P55903

b-depressantBotIT5


Na v:site4

LD50

1.1lgg�

1(B.germ

anica)102

P55904

b-depressantBotIT6


Na v:site4

LD50

0.1lgg�

1(B.germ

anica)282

P59864

b-depressantBaIT2

Buthacusarenicola

Na v:site4

LD50

3.5lgg�

1(B.germ

anica)283

P80962

TbIT-1

Tityusbahiensis

Na v:site4

LD50

4lg

g�1(M

.dom

estica)284

P60275

b-depressantBsIT1

Buthussindicus

Na v:site4

LD50

0.67lgg�

1(S.falculata)285

P82811

LD50

1.38lg

g�1(B.germ

anica)285

b-depressantBsIT2

Buthussindicus

Na v:site4

LD50

1.6lgg�

1(B.germ

anica)285

P82812

LD50

0.81lgg�

1(S.falculata)285

b-depressantBsIT3

Buthussindicus

Na v:site4

LD50

1.63lg

g�1(B.germ

anica)285

P82813

LD50

1.03lg

g�1(S.falculata)285

b-depressantBsIT4

Buthussindicus

Na v:site4

LD50

1.42lg

g�1(B.germ

anica)285

P82814

LD50

0.78lgg�

1(S.falculata)285

b-depressantLqhIT2

Leiurusquinquestriatushebraeus

Na v:site4

LD50

5.1lg

g�1(S.litura)114

Q26292

b-depressantLqqIT2

L.quinquestriatusquinquestriatus

Na v:site4

Kd

1.9nM

286

P19855

BoiTx1

(a-KTx3.10)

Buthusoccitanusisraelis

DrosophilaShaker

IC50

3.56

0.5nM

287

P0C908

BmBKTx1

(a-K

Tx19.1)

Mesobuthusmartensii

dSlo

IC50

194nM

(Drosophilamelanogaster)139

P83407

pSlo

IC50

82nM

(Periplanetaamericana)139

LaIT2(b-K

Tx-likepeptide)

Liochelesaustralasiae

ND

PD50

26lg

g�1(A

.dom

estica)152

C7G3K3

LaIT1

Liochelesaustralasiae

ND

LD50

1lg

/cricket(A

.dom

estica)152

P0C5F2

U1-liotoxin-Lw1a

Liocheleswaigiensis

ND

LD50

3.3lg

g�1(Tenebriomolitor)90

P0DJ08

LD50

5.4lgg�

1(A

.dom

estica)90

LD50

20.8lg

g�1(Luciliacuprina)90

Wasp

b-PMTX

Batozonellusmaculifrons

InsectNa v

channel

EC50

46lM

(para/tipE)120

P69392

Brh-I

Braconhebetor

ND

LD50

0.0023lg

g�1(G

alleria

mellonella)222

–

LD50

0.05lgg�

1(M

.sexta)222

LD50

0.033lg

g�1(Spodoptera

exigua)222

LD50

0.18lg

g�1(H

eliothisvirescens)222

LD50

0.045lg

g�1(H

eliothiszea)222

LD50

0.019lgg�

1(Trichoplusiani)222

Brh-V

Braconhebetor

ND

LD50

0.0001lg

g�1(G

.mellonella)222

–

LD50

0.04lgg�

1(M

.sexta)222

LD50

0.051lg

g�1(S.exigua)222

LD50

0.26lg

g�1(H

.virescens)222

LD50

0.085lg

g�1(H

.zea)222

LD50

0.0038lg

g�1(T.ni)222

Vespulakinin

Vespulamaculifrons

Bradikinin

receptors

IC50

90lM

(cockroach)231

P57672

Insecticidalpeptides

isolatedfrom

spider,scorpionandwaspvenomsarepresentedtogether

withtheirmoleculartargetandtheireffectivedose.Legend:

ND:Notdetermined.

TableI

ArthropodPeptides

withInsecticidalActivity(C

ontinued)

Arthropod

Peptide

Source

Target

Assay

Dose

UNIPROT

388 Schwartz et al.


SODIUM CHANNELS AS TARGETS FORINSECTICIDAL COMPOUNDSVoltage-gated sodium (Nav) channels are the molecular tar-

get of numerous therapeutic drugs (e.g., local anesthetics,

anticonvulsants and antiarrhythmics), neurotoxins and

chemical insecticides (e.g., pyrethroids, DDT, dihydropyra-

zoles, oxadiazines and N-alkylamides). These probes bind to

at least eight binding areas identified in Nav channels that are

important targets for agents used in insect control. Neuro-

toxin binding areas 1, 3, 4 and 6 are targets to peptide neuro-

toxins (see review45).

Site 1, located on the extracellular surface of the pore, and

composed of residues at the reentrant P-loops connecting S5

and S6 of all four domains, is the target of tetrodotoxin

(TTX), saxitoxin (STX) and l-conotoxins, blockers of Na+

conductance.46–50 The site 2, localized mainly at the S6 of DI

and DIV, recognizes the group of lipophilic activators batra-

chotoxin, veratridine, aconitine, and grayanotoxins. Breve-

toxins and ciguatoxins - lipophilic, cyclic polyether com-

pounds produced by marine dinoflagellates - target the neu-

rotoxin receptor site 5 located at DI S6.51,52 As result of

valuable neurotoxins structure-function studies conducted

recently by many groups, it has been proposed a revision on

the description of the binding sites 3 and 4.45 Neurotoxins

that target the main binding area located in the extracellular

loop between S3 and S4 in DIV (site 3 neurotoxins) slow or

inhibit the rapid inactivation of Nav channels by stabilizing

the voltage sensor S4 of domain IV in its deactivated posi-

tion.53,54 Among these neurotoxins are the classical scorpion

a-toxins, Type 1 and Type 2 sea anemone toxins, and various

spider toxins, including insecticidal toxins Tx4(6-1) and

Magi-1 to 4.55,56 Depending on specific amino acid residues,

the neurotoxins that target site 4 - located at DII, including

the extracellular linkers between S1–S3 and S3–S4 - can pro-

duce a hyperpolarizing shift in the activation (the classical

site 4 effect produced by scorpion b-toxins), or inhibit the

Na+ conductance, as lO-conotoxins and spider b-toxins(such as HWTX-IV).17,57,58 Finally, there are evidences sug-

gesting that a d-conotoxin (d-SVIE) interacts with a possible

site 6, constituted by conserved residues in the linker between

S3 and S4 of DIV, without the typical voltage-dependency of

the site 3 toxins.59 The performance of these toxins in the

field probably remains unaffected by the presence of pyreth-

roid resistance,10,60 so they show some potential for circum-

venting knockdown resistance to pyrethroids.

Two sodium channel genes highly homologous to the rat

brain sodium channel were shown in Drosophila mela-

nogaster brain61,62: DSCI gene, which is located on the right

arm of the second chromosome in proximity to the seizure

locus, and para gene, which overlaps with the paralysis locus

on the X chromosome. More recently, the functional expres-

sion in Xenopus oocytes confirmed that DSC1 encodes

indeed a Ca2+-selective cation channel.63 The para gene

shows 67% amino acid residue identity to the rat brain II so-

dium channel gene,57 showing the same degree to which the

vertebrate sodium channels resemble each other. It is fully

accepted that para and para-orthologs in other insect species

encode functional sodium channels.64–66 Although insect

and vertebrate sodium channels share high structural similar-

ities, and both are affected by the same classical pharmacol-

ogy probes, insect and vertebrate Na+-channels are indeed

distinguishable from a strictly pharmacological point of view,

and that a pharmacological distinction between insect and

noninsect Na+-channels occurred in the course of the long

coevolution of insects and their arachnid venomous preda-

tors.66,67

Insect-Selective Peptides of Arthropods Interacting

with Na+ Channels

Spider venom peptides targeting insect Nav channels charac-

terized to date can block the sodium conductance or slow

channel inactivation, depending of the main binding area,

validating them as potential insecticide targets (see review17).

Hainantoxin-I (HNTX-I) is the first insect-selective toxin

to selectively inhibit Na+ conductance.68 Among the spider

toxins acting as gating modifiers of inactivation, there are the

insect-selective Tx4(6-1)56,69,70 and Magi-2,55 while other in-

secticidal peptides, such as Magi-4, robustoxin, versutoxin,

d-HXTX-Hsp20.1, and Jingzhaotoxin-1 interact with both

vertebrate and invertebrate Nav channels.55,71–80 Although d-

palutoxins (d-PaluIT) modulate Nav channel inactivation,

similar to a-scorpion toxins, they were shown to compete

with insect b-scorpion toxins.81–83 Curtatoxins and l-aga-toxins, which are both specific to insects, also share an action

similar to classical b-toxins.9,84–86 Nevertheless, the number

of insect-selective spider toxins found to target Nav channels

is still limited, and several toxins are still awaiting eletrophy-

siological characterization of their target sites.

HNTX-I (or l-theraphotoxin-Hhn2b) is a 33-residue

peptide, with three disulfide-bonds, which displays a 15-fold

selectivity for the para/tipE insect Nav channel over the rat

Nav1.2/b1 channel, with no effect on other Nav channels,

being the first insect-selective blocker of Nav channels

reported.68 Contrary to many excitatory toxins that target

Nav channels, such as d-hexatoxins and l-agatoxins,87,88

HNTX-I has no effect on the inactivation kinetics. The three-







dimensional solution structure of HNTX-I belong to the In-

hibitory Cystine Knot (ICK) structural family and contain a

short triple-stranded antiparallel b-sheet (Figure 1A, PDB

code 1NIX). Structural comparison of this peptide with other

toxins suggests that the combination of the charged residues

and a vicinal hydrophobic patch should be responsible for

ligand binding.68

The Tx4(6-1) or d-ctenitoxin-Pn1a is a 48-residue long

peptide highly reticulated by five disulfide bonds isolated

from the venom of the Brazilian ‘‘armed’’ spider Phoneutria

nigriventer (Ctenidae). This peptide has Cys as the amino ter-

minal and carboxyl terminal residues of its primary

sequence,69,70 which is also characteristic of robustoxin (d-HXTX-Ar1a)71 and versutoxin (d-HXTX-Hv1a),72 from the

funnel-web spiders Atrax robustus and Hadronyche versuta,

respectively, which bind to site 3 of both insect and mammal

Nav channels.73–75,77,78,89,90 Tx4(6-1) is an insect-selective

toxin with a high degree of specificity toward different insect

species. It is highly toxic to house flies, causing excitatory

symptoms immediately after intrathoracical injection, and is

active against cockroaches (0.5–2.5 mg kg�1).69 It was shown

to be ineffective on mammalian Nav channels, but competed

with the a-like toxin 125I-Bom IV for binding on the site 3 of

insect Nav channel (IC50 around 25 nM), by slowing down

the sodium current inactivation.56

Corzo et al.55 isolated six neurotoxins from the Macrothele

gigas (Hexathelidae) spider venom, formerly named Magi-1

to Magi-6. Magi-1 (l-hexatoxin-Mg1b) had no effect when

injected to either mice (20 pmol g�1) or insects (32.8 nmol

g�1), although it seemed to bind to site 3 of the Nav channels

of cockroach synaptosomes. Magi-2 (l-HXTX-Mg1a), Magi-

3 (l-HXTX-Mg2a), Magi-4 (d-HXTX-Mg1a), Magi-5 (b-HXTX-Mg1a), and Magi-6 (U7-HXTX-Mg1a) caused paral-

ysis to lepidopteran larvae (LD50: 17.6, 10.3, 1.2, 8.6, and 3.1

FIGURE 1 Examples of structural diversity of molecules with insecticidal activity isolated from

Arthropod venoms. Schematic tridimensional structure view of arthropod insecticidal peptides act-

ing on sodium channels (A), potassium channels (B), calcium channels (C), or unidentified targets

(D). PDB codes: 1NIX for HNTX-I, a Na+ channel toxin isolated from Haplopelma hainanum

spider; 1V90 for d-PaluIT1 and 1V91 for d-PaluIT2, two insecticidal Na+ channel toxins from the

spider Pireneitega luctuosa; 1G92 for Poneratoxin, from Paraponera clavata ant; 1LQH, for LqhaIT,an insecticidal a-NaScTx isolated from the scorpion Leiurus quinquestriatus hebraeus that binds to

site 3 of domain IV; 1WWN, for BmKIT1, an excitatory scorpion toxin from Mesobuthus martensii

Karsch; 1DL0, for J-ACTX-Hv1c, a BKCa channel blocker from Hadronyche versuta spider venom;

2CRD for charybdotoxin, a potent selective inhibitor of BKCa channels from L. quinquestriatus

hebraeus; 1AXH for x-ACTX-Hv1a, a blocker of M-LVA and HVA Cav channels from H. versuta;

1G9P from Hadronyche versuta, an inhibitor of insect Cav channel; 1I25 Huwentoxin-II, an insect

paralyzing toxin from Haplopelma hainanum; 2LDS for LaIT1, a short-chain peptide from the

venom of scorpion Liocheles australasiae. Structures were prepared using OpenAstexViewer avail-

able at http://www.ebi.ac.uk/pdbe/. The structures were colored by domain following Olderado,

which provides analysis of clustering and domain composition for NMR structure ensembles

(http://www.ebi.ac.uk/pdbe-apps/nmr/olderado/main.html).

390 Schwartz et al.


nmol g�1, respectively), although only Magi-2 and 3 were

not toxic to mice at 20 pmol g�1. Magi-1, 2, 3, 4 and 5 seem

to bind to site 3 of the Nav channels of cockroach synapto-

somes, Magi-4 being the one that exhibited the strongest

competition against LqhaIT for site 3 of insect Nav channels

(Ki ¼ 0.05 nM). None of these toxins competed with any

toxin in binding to rat brain synaptosomes. Only Magi-5

competed with both the b-scorpion toxin CssIV, that binds

to site 4 of the rat brain sodium channel, and with the insect

sodium a-toxin LqhaIT.55

The d-HXTX-Hsp20.1a, from the funnel-web spider Illa-

wara wisharti, slows the inactivation of vertebrate TTX-sensi-

tive and insect Nav channels.17,76 The Jingzhaotoxin-1 (d-TRTX-Cj1a), from the tarantula Chilobracys jingzhao,79 is

also suggested to inhibit sodium channel inactivation of ver-

tebrate (Nav1.5, Nav1.6, and Nav1.7) and invertebrate Navchannels,80,91 besides promoting very weak inhibition of

voltage-gated potassium (Kv) channels of the subtype

Kv2.1.92

The d-PaluITs, l-agatoxins (l-Aga), and curtatoxins are

highly homologous peptides composed by 36–37 amino acid

residues and cross-linked by four disulfide bridges forming

an ICK motif.17 The d-palutoxins IT1 to IT4 (or d-AMATX-

PI1a to PI1d) are insect-selective Nav-channel toxins isolated

from the Pireneitega luctuosa (Amaurobiidae) spider

venom.81 These four peptides were significantly toxic against

larvae of the crop pest Spodoptera litura (Lepidoptera), the d-PaluIT1 being the most active toxin. Only d-PaluIT2 was

toxic to mice at a dose of 2 lg per animal.81 They were

shown to slow the inactivation of inward Na+ currents in

cockroach axons in a way similar to the insecticidal scorpion

a-toxins,81 although they displace the site 4 excitatory scor-

pion b-toxin Bj-xtrIT from binding on cockroach mem-

branes and fail to displace LqhaIT binding.82 These toxins

also have no effect on Nav1.2/b1 channels at concentrations

up to 10 lM.83 The structures of d-paluIT1 and d-paluIT2(Figure 1A, PDB codes 1V90 and 1V91) contain respectively

two- and three-stranded antiparallel b-sheets as unique sec-

ondary structure and the recognition of insect Nav channels

by these toxins involves the b-sheet, in addition to loops I

and IV.86 Similar to d-paluIT1, the structure of l-Aga-I isalso formed by two-stranded antiparallel b-sheets (PDB code

1EIT).88

The l-agatoxins l-Aga-I to VI (or l-AGTX-Aa1a to

Aa1f), from the venom of Agelenopsis aperta (Agelenidae),

constitute a family of peptides that affect only insect Navchannels.84 These six peptides promote a gradual but irre-

versible paralysis in lepidopterous insects.84 Electrophysio-

logical studies show that these toxins modify presynaptic Navchannels, causing them to open at the normal resting poten-

tial of the nerve, which leads to repetitive firing in the termi-

nal branches of insect motor axons and spontaneous release

of neurotransmitter.85 Although more studies are required to

confirm its site of action, the curtatoxins I to III (l-AGTX-Hc1a to Hc1c), from the venom of Hololena curta (Agene-

lidae), seem to have a similar mode of action, and produced

an immediate and irreversible paralysis in the cricket A.

domestica.9,86

The scorpion sodium channel modulators (NaScTxs) are

long-chain peptides with 55–76 amino acid residues and

cross-linked by three or four disulfide bridges.93,94 The

NaScTxs can be classified in two families, a and b, accordingto the mode of action on Na+ channels.16 The a-NaScTxsbind to site 3 of domain IV, slowing the inactivation of the

channel. According to structural reasons and the specificity

of interaction with mammal or insect Na+ channels, they

were divided into three subgroups: (a) classical a-toxins arehighly toxic to mammals, such as AaHII,95 from Androctonus

australis Hector and LqhII96 from Leiurus quinquestriatus

hebraeus; (b) anti-insect a-toxins, which are highly toxic to

insects19; (c) a-like toxins, which act on both mammal and

insect Na+ channels (for review see18,19).

The toxins LqhaIT and Lqh III, from Leiurus quinquestria-

tus hebraeus, are anti-insect a- and a-like toxins, respec-

tively.96–99 Lqh III acts on both mammal and insect Na+

channels,96 while LqhaIT presents a weaker toxicity against

mammals, with a LD50 value of 5 mg kg�1 by subcutaneous

injection in mice.97 LqhaIT is a 64 amino acid residue pep-

tide, containing an a-helix and a three stranded antiparallel

b-sheet (Figure 1A, PDB code 1LQH).100 Site-directed muta-

genesis conducted in the C-terminal domain of LqhaITreduced antimammalian toxicity without significantly affect-

ing insecticidal potency.101 Another anti-insect a-toxin,BotIT1, a 64 amino acid residue long peptide purified from

Buthus occitanus tunetanus, was able to slow down the inacti-

vation mechanism of sodium channels on the cockroach

axon.102 BjaIT, an insect-selective a-toxin isolated from

Buthotus judaicus, inhibits the inactivation mechanism of

para/TipE channel (100 nM) without affecting Nav1.2/b1channel.103

The b-NaScTxs bind to site 4 of domain II, shifting the

voltage activation of the channel to more negative membrane

potentials.16,104 The b-toxins are subdivided into four subfa-

milies18,104,105: (a) antimammal b-NaScTxs, which are highly

toxic to mammals, and modulate the activation of Na+ chan-

nels in mammalian brain, such as Cn2 from Centruroides

noxius106,107 and Css4 from C. sufussus sufussus108, this group







of toxins was isolated mainly from scorpions of the New

World109; (b) b-excitatory anti-insect, which are harmless to

mammals, and induce spastic paralysis, caused by repetitive

activity of motor nerves, resulting from activation of Na+

channels to more negative membrane potentials.110 The con-

sequence of this mode of action is a contracture on the skele-

tal musculature, followed by a muscle fatigue, which is re-

sponsible for the paralysis.19 Examples are AahIT1 from

Androctonus australis Hector,111 and BmKIT1,112 from Meso-

buthus martensii, which is a 70 amino acid residue toxin,

with two a-helix anchored against a three-stranded b-sheetstabilized by three disulfide bonds113 (Figure 1A, PDB code

1WWN); (c) b-depressant anti-insect toxins that induce flac-cid paralysis to insects, induced by a short transient phase of

contractility in larvae,19 such as LqhIT2114 from Leiurus

quinquestriatus hebraeus; (d) b-like toxins that are highly

active on mammals and insects, like Lqhb1 from Leiurus

quinquestriatus hebraeus, which competes with anti-insect

and antimammalian b-toxins in binding to cockroach and

rat brain synaptosomes, respectively.104 BmK IT2, a depres-

sant insect-selective toxin isolated from Mesobuthus martensii

Karsch115,116 is able to bind specifically to insect sodium

channels,117 interacting to two binding regions, comprising

domains II and III of DmNav1.118 When compared with the

a-insect peptides, the number of b-insect toxins is almost

five times greater. Among these b-toxins, *65% of them are

b-insect depressant toxins. Until recently, these b-anti-insectNaScTxs were restricted to Old World scorpions. The first

anti-insect excitatory b-class NaScTx from New World was

described in Tityus pachyurus scorpion venom.119 More

examples of a and b anti-insect NaScTxs are shown in Table

I.

Even though studies on sodium channel modulators from

hymenopteran venoms are still scarce, these peptides would

provide advantages for studying Nav channels function as

they are devoid of disulfide bridges and shorter than those

from other arthropod groups.

In recent decades, a class with great insecticide potential

was isolated from solitary wasps, the pompilidotoxins.

a-Pompilidotoxin (a-PMTX) and b-pompilidotoxin

(b-PMTX) are the members of this class constituting two

small peptides with 13 amino acid residues purified from the

venom of the solitary spider wasps Anoplius samariensis and

Batozonellus maculifrons, respectively. Differently from scor-

pion and anemones, a- and b-PMTXs act on insect voltage-

gated sodium channels (DmNav1) increasing the steady-state

current component without any increase in the slow compo-

nent of the sodium inactivating current. Moreover, the dose-

dependent increase in this steady-state component was corre-

lated with the dose-dependent decrease in the fast compo-

nent.120

The tropical predatory ant Paraponera clavata contains a

potent insect-specific neurotoxin named poneratoxin. This

toxin contains 25 amino acid residues, folded as two a-heli-ces connect by a b-turn (Figure 1A, PDB code 1G92). One of

the helices is apolar, whereas the second contains polar and

charged amino acid residues.44 When injected in Spodoptera

frugiperda larvae, synthetic poneratoxin caused reversible pa-

ralysis 2 min after injection.44 Poneratoxins affect the volt-

age-dependent sodium channels and block nicotinic synaptic

transmission in insect central nervous system in a concentra-

tion-dependent manner.37

POTASSIUM CHANNELS AS TARGETS FORINSECTICIDAL COMPOUNDSSince the 70’s efforts have been made to develop selective

insecticides that act specifically on biochemical routes that

are present in particular insect groups but with properties

that differ from other insecticides.121 Consequently, insect

growth regulator (IGR) insecticides were developed, such as

the ecdysone agonists.122 One example of these ecdysone

agonists are the 1,2-dibenzoyl-1-tert-butylhydrazines. Its

prototype, RH-5849, preferentially blocks the late sustained

component (IK) of the potassium current with little effect on

the initial transient component (IA).123

Potassium channels have the highest diversity between ion

channels and can be grouped into four families: the voltage-

gated channels (Kv, with 12 subfamilies), the calcium-acti-

vated ones (KCa, 3 subfamilies), the inward-rectifiers (Kir,

with 7 subfamilies), and those with two tandem-pores (K2P,

with 16 subfamilies).124 With great relevance for the study of

new insecticides from the venom of arthropods, the calcium-

activated potassium channels (KCa) are a heterogeneous fam-

ily of potassium channels whose open probability is increased

by the elevation of cytosolic calcium to cause membrane hy-

perpolarization. They can be found in a wide range of differ-

ent tissues and organisms and can be divided into three sub-

families: BKCa or large (big)-conductance KCa channels, SKCa

or small-conductance KCa channels and also IKCa or interme-

diate-conductance KCa channels.125

Large-conductance KCa channels, also termed BKCa

(KCa1.1), Maxi-K or Slo1 channels, are activated by mem-

brane depolarization and by increases in intracellular Ca2+

concentration.126 They were first studied in smooth muscle

cells, where they are key players in setting the contractile

tone, but they are also abundant in other tissues, such as

brain, pancreas, and urinary bladder. BKCa channels are pres-

ent in many multicellular organisms, such as nematodes,

392 Schwartz et al.


insects, and mammals.127 These channels play an important

role in many cellular functions such as action potential repo-

larization, neuronal excitability, neurotransmitter release,

hormone secretion, tuning of cochlear hair cells, innate im-

munity, and modulation of the tone of vascular, airway, uter-

ine, gastrointestinal, and urinary bladder smooth muscle tis-

sues. Thus, new activators and blockers of BKCa channels

could help understand the physiology of these channels and

also have applications as neuroprotectors or as therapeutics

in certain disease states.127 In addition, the discovery of novel

insect-selective ligands of these channels could provide new

tools for biopesticide engineering.128


with K+ Channels

The Janus-faced atracotoxins (J-ACTXs) or j-hexatoxins area family of insect-selective neurotoxins from the venom of

the Australian funnel-web spider Hadronyche versuta (Hexa-

thelidae), each with 36 or 37 amino acid residues and four

disulfide bridges.129 The three-dimensional structure of J-

ACTX-Hv1c (the most potent of the J-ACTXs) revealed a

rare vicinal disulfide bridge, connecting Cys13 to Cys14,

which was demonstrated to be critical for insecticidal activ-

ity. The other three central disulfide bonds form an ICK

motif with two-stranded antiparallel b-sheets (Figure 1B,

PDB code 1DL0).129 All peptides from this family are toxic

to house cricket, flies, mealworms, and budworms, with no

effects on mice, chicken and rat preparations.129–132 In cock-

roach dorsal unpaired median (DUM) neuron preparation,

it was shown that J-ACTX-Hv1c is a high-affinity blocker of

insect BKCa currents, with minimal effects on mouse or rat

BKCa channels and no effect on Cav and Nav channel currents

at 1 lM concentration.128 The amino acid variations in the

pore region of BKCa channels appear to be sufficient to

explain the insect selectivity of J-ACTX-Hv1c, which is most

likely a channel blocker than a gating modifier.128 Due to the

870-fold higher selectivity of J-ACTX-Hv1c for insect BKCa

channels than charybdotoxin and to the small molecular epi-

tope on this toxin that mediates its interaction with these

channels, comprising only five spatially proximal resi-

dues,128,131 the study of this epitope should provide an acces-

sible template for the rational design of small-molecule

insecticides that specifically target insect BKCa channels.

Another group of spider toxins that specifically targets

BKCa channels is composed of three new ‘‘short-loop’’ inhibi-

tory cysteine knot insecticidal toxins named j-theraphotox-

ins: j-TRTX-Ec2a, j-TRTX-Ec2b, and j-TRTX-Ec2c.133

These three 29-residue peptides, isolated from the venom of

the East African tarantula Eurocratoscelus constrictus, were

found to be high-affinity blockers of insect BKCa channel cur-

rents (IC50 ¼ 3–25 nM), displaying no significantly effect on

other insect Kv, Nav, or both M-LVA and HVA Cav channels

on cockroach DUM neurons. j-TRTX-Ec2a and j-TRTX-Ec2b presented insect-selective effects, whereas j-TRTX-Ec2cwas also toxic to mice when injected intracerebroventricu-

larly. Differently to J-ACTX-Hv1c, these toxins are thought

to interact with the turret and/or loop region of the external

entrance to the channel and do not project deeply into the

channel pore.133 Along with J-ACTX-Hv1c, the jTRTX-Ec2toxins validate insect BKCa channels as potential targets for

insecticides.

Charybdotoxin (a-KTx 1.1),134 isolated from Leiurus

quinquestriatus hebraeus (Yellow scorpion), is a potent selec-

tive inhibitor of BKCa channels and is composed of 37 amino

acid residues, forming a triple-stranded beta-sheet and an a -

helix (Figure 1B, PDB code 2CRD).135 Iberiotoxin (IbTx, a-KTx 1.3), a toxin isolated from the venom of the scorpion

Mesobuthus tumulus, is a selective inhibitor of BKCa chan-

nels.136,137 Charybdotoxin and IbTx were able to block pSlo,

the BKCa channel a subunit of the cockroach Periplaneta

americana. These channels are found in the octopaminergic

DUM neurons and peptidergic midline neurons in Peripla-

neta abdominal ganglia.138

BmBKTx1 (a-KTx 19.1) is a 31 amino acid residue long

toxin purified from the venom of the Asian scorpion Meso-

buthus martensii Karsch that presented a blocking effect on

pSlo and dSlo, the a subunit of BK channels of the fruit fly,

but hardly affected hSlo (a subunit of human BK channels)

currents, even at concentrations as high as 10 lM, suggesting

that the toxin might be insect specific.139

Many a-KTxs have been proved against the insect Kv1

channel, the Shaker channel. Some examples are charybdo-

toxin,140 MeuKTX141 (a-KTx 3) from the venom of the scor-

pion Mesobuthus eupeus, HgeTx1142 (a-KTx 6.14) from

Hadrurus gertschi, TdK2143 (a-KTx 18.2) from Tityus discrep-

ans, Tco30144 (a-KTx 12.3) from Tityus costatus, Pi1145 (a-KTx 6.1) and Pi4146 (a-KTx 6.4) from Pandinus imperator,

Tc1147 (a-KTx 13.1) from Tityus obscurus, agitoxin 1 (a-KTx3.4), 2 (a-KTx 3.2), and 3 (a-KTx 3.3)148 from Leiurus quin-

questriatus hebraeus. Despite the variety of a-KTxs so far

described, few of them showed specificity to insect Kv1 chan-

nels (Shaker) with no or low activity in mammal Kv1 chan-

nels. An example is the toxin BoiTx1 (a-KTx 3.10) purified

from Buthus occitanus israelis, a 37 amino acid-long peptide

containing six conserved cysteines. This peptide inhibited







currents through Drosophila Shaker channels, but it had a

much lesser effect on their mammalian orthologs.149

Beta-KTx, or scorpine-like peptides, or even ‘‘orphan pep-

tides,’’ are long chain peptides containing three disulfide

bridges with two recognizable domains: a freely moving N-

terminal amino acid sequence and a tightly folded C-termi-

nal region with a cysteine-stabilized alpha/beta (CS-ab)motif. The full-length scorpine-like peptides can lyse cells or

kill microorganisms, while the C-terminal domain contains

the CS-ab motif that can block K+ channels.150 From the

venom of the Thai giant scorpion, Heterometrus laoticus, it

was purified the peptide named Heteroscorpine-1, which

present high similarity to other scorpine-like peptides, and

was able to paralyze crickets.151

LaIT2 was isolated from Liocheles australasiae (Hemiscor-

piidae), is composed of 59 amino acids with three disulfide

bridges, and shares sequence similarity with the scorpion b-KTx peptides.152 Its six-cysteine residues are conserved when

compared with the rest of the b-KTxs peptides. These mole-

cules are known as orphan peptides because of the uncer-

tainty of their target, and they were divided into three subfa-

milies: b-KTx from Buthidae, scorpine-like peptides and

peptides similar to BmTXKb2.153 LaIT2 showed 46% identity

with the HgebKTx,154 belonging to the subfamily similar to

BmTXKb2. LaIT2 showed a paralytic effect in Acheta domes-

tica and produced evidence of an antimicrobial effect.152 The

insecticidal activity of a b-KTx peptide gives rise to a new

approach in the exploration of insecticide toxins.

Studies describing the action of wasp, bee, ant, and centi-

pede venom peptides on insect potassium channels and the

trials to evaluate their insecticidal activity have not been

reported so far. Two potassium channel blockers were iso-

lated from bee venoms, named apamin and tertiapin.155,156

Apamin is a very potent and highly selective antagonist of

all three subtypes of SKCa-channel and has been largely used

to study their function in many cell types, including the

physiological mechanisms involved in higher brain functions,

especially cognitive processes or in the control of mood in

mammals.157–159

Tertiapin blocks two different types of potassium chan-

nels, namely the inward rectifier potassium channels (Kir)

and BKCa channels. This peptide binds specifically to differ-

ent subunits of the Kir channel with nanomolar affinities,

namely GIRK1 (Kir 3.1), GIRK4 (Kir 3.4), and ROMK1 (Kir

1.1), inducing a dose-dependent block of the potassium cur-

rent.160

Apamin and tertiapin are very useful tools for studying

the physiology and the structure-function relationship of

these channels. They might also be powerful ligands for puri-

fying functional channels as well as for screening pharmaceu-

tical agents against these channels.

CALCIUM CHANNELS AS TARGETS FORINSECTICIDAL COMPOUNDSElevation of intracellular Ca2+ concentration ([Ca2+]i) ini-

tiates a cascade of signal transduction events that culminate

in a variety of physiological processes including secretion of

neurotransmitters, activation of transcription factors, muscle

contraction, and many others.161 In mammals, reptiles,

insects, and other animal taxa, the depolarization of the

external membrane of cardiac and skeletal muscle cells acti-

vates voltage-dependent Ca2+ channels, which in turn acti-

vate Ca2+ release channel/ryanodine receptors (RyRs) of the

sarcoplasmic reticulum (SR) to elevate [Ca2+]i and initiate

myofilament contraction. The release of Ca2+ by RyRs is so

abrupt and massive that an intrinsic inactivation process is

safely in place to avoid saturating cytosolic compartments

with Ca2+ and subsequent contractile paralysis.162 Indeed, ry-

anodine, a plant alkaloid and an important ligand used to

characterize and purify receptors, has served as a natural bo-

tanical insecticide, causing muscle paralysis and death by

opening RyRs with high potency.163 The insecticides Rynaxy-

pyr and Cyazypyr present inherent selectivity to insect RyRs

over their mammalian counterparts (for a review see164). The

future prospects for RyRs as a commercially validated target

site for insect control chemicals should be considered.165

Insects have a more limited voltage activated calcium

channel (Cav) repertoire than vertebrates (see review43).

While the human genome encodes 10 a1 subunits (the pore-

forming unit that determines the pharmacology of the chan-

nel), four b subunits, four a2-d complexes, and seven c subu-nits that constitute the seven Cav channels, the genome of

Drosophila melanogaster, for example, encodes only three a1subunits, a b subunit, three a2-d complexes, and possibly

one c subunit.166,167 These three a1 subunits from D. mela-

nogaster, Dmca1D, Dmca1A, and Ca-a1T, have no more than

68% identity with mammalian a1 subunits, and each one of

these insect Cav channels is orthologous to a single mammal

subfamily, Cav1, Cav2, and Cav3, respectively.43 Despite this

moderate homology, these channels have different electro-

physiological properties and different sensitivities to various

pharmacological agents, and therefore would not be suitable

to receive the same classification.43 Although their inventory

is likely to be increased by alternative splicing of Cav channel

transcripts and RNA editing,168,169 the small repertoire of

Cav-a1 genes in insects suggests that these channels play an

essential physiological role.170,171

394 Schwartz et al.



with Ca2+ Channels

Among the spider toxins that interact with Cav channels, the

x-atracotoxin-Hv1a (x-ACTX-Hv1a),172–176 x-ACTX-Ar1a,177 x-ACTX-Hv2a,178 and the huwentoxin-V (HWTX-

V)179,180 are examples of insect-selective peptides toxic to a

wide variety of insects and with no effects on mammals.

Other toxins, such as CSTX-1 (x-ctenitoxin-Cs1a)181–183

and those belonging to the x-agatoxin family (see review184),

are taxonomically promiscuous, acting on both vertebrate

and invertebrate Cav channels. Although more studies are

required, PLTX-II (x-plectoxin Pt1a) also appears to be spe-

cific for insect Cav channels.185–187

The x-ACTX-Hv1a or x-hexatoxin-Hv1a, isolated from

the venom of Australian funnel-web spider Hadronyche ver-

suta (Hexathelidae) is a peptide of 37 amino acid residues

and 3 disulfide bonds that form an ICK motif. Its structure

consists of a solvent-accessible b-hairpin at the C-terminal

protruding from a disulfide-rich globular core comprising

four b-turns (Figure 1C, PDB code 1AXH).172 This toxin is

considered one of the most potent insecticidal peptides char-

acterized to date, and is toxic to a variety of arthropods of ag-

ricultural importance belonging to the Coleoptera, Lepidop-

tera, and Diptera orders, with no reported effects on mam-

mals.172,174 In insect preparations, the x-ACTX-Hv1a acts

directly on CNS neurons and not on interganglionar axons

or peripheral neuromuscular junctions.175

Chong et al.177 demonstrated that x-ACTX-Hv1a and x-ACTX-Ar1a reversibly blocked M-LVA current (mid-low)

and HVA (high-voltage activated) Cav channels in cockroach

DUM. These toxins appear to act as pore blockers, since they

do not affect the kinetics of activation or inactivation and are

voltage independent. x-ACTX-Hv1a had no effect on heter-

ologous mice channels Cav1.2 (L type), Cav2.1 (P/Q), and

Cav2.2 (N-type).173,174 The x-ACTX-Ar1a, a 37 amino acid

residue long peptide isolated from the Sydney funnel-web

spider Atrax robutus,177 induced excitatory symptoms, fol-

lowed by flaccid paralysis and death in crickets, but showed

no effect on vertebrate smooth and skeletal muscle neuro-

muscular preparations in concentrations up to 1 lM. Also in

DUM neurons, x-ACTX-Ar1a (1 lM) had no effect on po-

tassium channels, and blocked 18 6 5% the peak current of

Na+ in Nav channels, proving to be more effective in Cavchannels.177

The x-ACTX-Hv2a, a prototype member of the x-ACTX-2 family, which includes insecticidal peptides insecticides com-

posed of 42–45 amino acid residues, caused immediate and

nonreversible paralysis when injected into crickets.178 This

same toxin, when injected into the brain neurons of bees,

inhibited HVA current type of Cav channels with EC50 of

around 130 pM. The x-ACTX-Hv2a did not inhibit TTX-sen-

sitive sodium currents (INa) and potassium currents (IK) in

bee brain neurons, and did not significantly alter INa in the

sensory neurons of mice.178 The disulfide-rich region of x-ACTX-Hv2a is organized into a compact globular domain

containing a small 310-helix and a short b-hairpin. In contrast

to the highly ordered globular core, the entire lipophilic C-ter-

minal tail is disordered in solution, which is essential for chan-

nel blocking activity (Figure 1C, PDB code 1G9P).178

The HWTX-V (x-TRTX-Hh2a) from the Chinese bird

spider Haplopelma schmidti, is a peptide with 35 amino acid

residues first sequenced from its precursor sequence,188 and

which was isolated from the venom together with its natural

mutant mHWTX-V.179 The HWTX-V can reversibly paralyze

locusts and cockroaches for several hours, and cause death at

higher doses. Its natural mutant, however, showed no signifi-

cant effect on locust and cockroaches, indicating that the last

C-terminal residues (Phe34 and Ser35), which are absent in

this mutant, are essential for biological activity.179 In whole-

cell patch-clamp configuration, HWTX-V specifically inhib-

ited high-voltage-activated (HVA) calcium channels in adult

cockroach DUM neurons (IC50 ¼ 219 nM), with no evident

effects on LVA currents from both rat dorsal root ganglion

and cockroach DUM neurons and only a weak inhibition on

mammalian neuronal HVA calcium currents at 1 lM concen-

tration. At this same concentration, no effect was detected on

Kv and Nav channels.180 This toxin had also no effect on mice

by intraabdominal or intracerebroventricular injection,179

making the HWTX-Van interesting tool for the development

of new insecticidal compounds.

The toxins of the family of x-agatoxins (x-Aga), from the

venom of the spider Agelenopsis aperta (see review184), are

shown to act on both vertebrate and invertebrate Cav chan-

nels. The x-Aga-IVA (x-AGTX-Aa4a), known as the proto-

type vertebrate P/Q type Cav channel (Cav2.1) antagonist,189

presented an effect similar to the x-ACTX-Hv2a in bee neu-

rons, inhibiting current type HVA Cav channels, but with an

EC50 (10 nM) significantly higher than that of x-ACTX-Hv2a.178 While high concentrations (1 lM) of x-ACTX-HV2a had little effect on sensory neurons of mice, the x-Aga-IVA inhibited ICa currents with EC50–20 nM in the same

assay.

The CSTX-1 is another insecticidal peptide that targets

both vertebrate and invertebrate Cav channels. Being the

most abundant toxin in the venom of the spider Cupiennius

salei (Ctenidae),181 it specifically blocks both mid/low (M-

LVA) and HVA insect Cav channels in cockroach DUM neu-







rons, as well as L-type Cav channels (Cav1) in rat neurons,

with no effects on other voltage-gated ion currents of rat

neurons, such as INa or IK.183 It was demonstrated that the

insecticidal activity of this peptide could be increased by syn-

ergistic interactions with different components from C. salei

venom, thereby proving the importance of these synergistic

strategies to the enhancement of venom efficacy during prey

envenomation.181,190,191

The PLTX-II, a peptide of 44 amino acid residues long

peptide, isolated from the venom of the spider Plectreurys

tristes,185 has at its C-terminal end a rare post-translational

modification, an O-palmitoyl threonine amino acid residue,

essential for its action.192 This toxin blocks Cav channels in

presynaptic nerve terminals of Drosophila,185 being the

Dmca1a Cav channels its probable molecular target,186 what,

due to its neuronal location, may explain the ineffectiveness

of the toxin on Drosophila muscle.187 The effects of PLTX-II

have not been much explored on Cav channels of vertebrates

yet, but the toxin did not block the neuromuscular junction

of amphibians (frogs) at a concentration of 10�8M.187

There are no Ca2+ channel insect-selective toxins

described to scorpions to date. However, some scorpion tox-

ins can be used as important tools to the study of Ca2+ chan-

nels, as well as insecticide prototypes. Purified from the

venom of the African scorpion Pandinus imperator, impera-

toxin A (IpTxa) was the first scorpion toxin that demon-

strated capacity to bind to RyRs with high affinity and speci-

ficity.193 IpTxa is a highly basic, short (33-amino acid) pep-

tide stabilized by three disulphide bridges and folding along

an inhibitor cystine-knot (ICK) motif.194,195 Later on, from

Scorpio maurus palmatus scorpion venom, the maurocalcin

(MCa) was classified based on sequence similarity to

IpTxa.196 It also presents amphipathicity, which confers the

capacity of these peptides to penetrate external mem-

branes.197,198 Other scorpion peptides with high sequence

similarity to IpTxa and MCa were subsequently described:

opicalcin 1 and 2,199 hemicalcin,200 and hadrucalcine.201

GLUTAMATERGIC NEUROTRANSMISSIONGlutamate is the main mediator of excitatory signals in the

mammalian brain and spinal cord. In invertebrates, in addi-

tion to its importance as CNS neurotransmitter, glutamate is

the main transmitter at the neuromuscular junction, assum-

ing both excitatory and inhibitory functions. The excitatory

or depolarizing response is mediated by a cation (mainly

Na+) selective channel, whereas the inhibitory or hyperpola-

rizing response is mediated by Cl� selective channel, both

found in the insect CNS and neuromuscular junction.202

The glutamate receptors are categorized according to their

differential intracellular signal transduction mechanisms and to

the molecular homologies in: (a) metabotropic receptors

(mGluRs) that mediate their function via intracellular G-pro-

teins; (b) ionotropic receptors (iGluRs), which are cation selec-

tive channels and are subclassified into N-methyl-D-aspartate

(NMDA), a-amino-3-hydroxy-5-methyl-4-isoxazale propionic

acid (AMPA, also named quisqualate receptor), and kainate

receptors.203,204 After its action, glutamate is removed from the

synaptic cleft by high-affinity transporters that are embedded

in the membrane of neuronal and glial cells and are typically

dependent on the Na+ gradient.205 Considering that there is no

extracellular enzyme that metabolizes glutamate to any signifi-

cant degree,205 the only rapid way to remove glutamate from

the synaptic cleft is by cellular uptake.206,207 Complementarily,

neurotransmitter transporters responsible for this uptake are

involved in the modulation of electrical signals, the transport

of water and ions, and it has been proposed that they play a

dynamic role in the control of CNS excitability.208,209

Insects and crustaceans neuromuscular preparations were

used in the past as model systems to study glutamate recep-

tor pharmacology. In general, glutamate receptors of insect

neuromuscular junction are similar to the mammalian

AMPA/kainate subtype. The venoms of arthropods contain

peptides that produce paralysis when injected into insects

and behave as modulators of glutamate transmission when

applied in neuromuscular preparations.210,211

As far as we know, there is no arthropod venom peptide ca-

pable to interact to glutamate receptors up to date. However,

there are plenty of reports focused on describing peptides that

interfere with the release or uptake of glutamate. The next sec-

tion will present a glimpse on some insect-selective arthropod

venom peptides that affect the glutamatergic neurotransmission.

Insect-Selective Peptides of Arthropods that Affect

Glutamatergic Neurotransmission

The spider P. nigriventer peptide fraction 4 (PhTx4)69 inhib-

its glutamate uptake in a dose-response manner in rat corti-

cal synaptosomes (IC50 ¼ 2.35 6 0.9 lg mL�1). It caused pa-

ralysis in house flies when injected intrathoracically and was

only toxic to mice at doses higher than 9.5 lg/animal by i.c.v.

injections.212 This venom fraction is composed of seven tox-

ins, referred to as P. nigriventer peptide toxins 4(1–7)

[PnTx4(1–7)]. Among them, P. nigriventer peptide toxins

4(3–7) also inhibit glutamate uptake.212

Isolated from this fraction, P. nigriventer peptide toxin 4-3

(PnTx4-3 or d-ctenitoxin-Pn1b) is a 48-residue long peptide

that has been reported to decrease glutamate release from rat

brain synaptosomes.213 This toxin induces immediate excita-

396 Schwartz et al.


tory effects when injected intrathoracically into house flies

and cockroaches, with no apparent signs of intoxication in

mice (30 lg/mice via i.c.v.).213

Also from the PhTx4 fraction, the P. nigriventer peptide

toxin 4(5–5) [PnTx4(5–5) or c-ctenitoxin-Pn1a] is a single-

chain insecticidal molecule composed of 47 amino acid resi-

dues, including 10 cysteines, which reversibly inhibits

NMDA receptor-generated currents in rat hippocampal neu-

rons. This toxin is highly toxic to the housefly (Musca domes-

tica), cockroach (Periplaneta americana), and cricket (Acheta

domestica), with no apparent macroscopic effects when

injected via i.c.v. (30 lg) into mice.214

As described on section ‘‘Insect-selective peptides of

arthropods interacting with Na+ channels,’’ the spider toxin

Tx4(6-1), which is an insect-selective Na+ channel toxin

highly toxic to house flies and cockroaches and not toxic to

mice (30 lg by i.c.v. injection),69 also affects the peripheral

nervous system of insects by stimulating glutamate release at

the neuromuscular junction.56

Recently, several studies have demonstrated the effect of

some a- and b- Na+ channel scorpion toxins on the gluta-

mate release. Tityus serrulatus scorpion Ts7 (UniProtKB

P15226), a b-like NaScTx, was shown to evoke glutamate

release in an incremental, dose - dependent manner.215 The

results suggested that this peptide preferentially binds to Na+

channels close to the active zones for glutamate release and

indicated that modifications of the activation or inactivation

of Na+-channels can lead to very different changes in the cy-

tosolic concentrations of free Na+ and Ca2+, with consequen-

ces for neurotransmission.215

Chromatographic fractions of Mesobuthus tamulus scor-

pion venom, showing toxicity to Lepidoptera larvae and

mice, inhibited the activity of the enzyme glutamate dehy-

drogenase, which is related to the glutamate metabolism.216

The venom component responsible to affect the enzyme ac-

tivity remains to be clarified.

Several parasitoid wasps that belong to the families Pompi-

lidae, Sphecidae, Mutillidae, and Bethylidae use their venoms

to paralyze spiders or larvae of insects.217 Their venoms act

slowly and cause a flaccid paralysis, part of this effect being

related to the blockage of glutamatergic neurotransmission.

The venom of the parasitic wasp Bracon hebetor has been

known to act against lepidopterous larvae for over 50 years.

The action of the venom is specie-specific, as it is extremely

active in Lepidoptera, less active in honeybee workers, much

less active in locusts and mealworms and probably inactive in

noninsects. This venom blocks neuromuscular transmission

presynaptically in Philosamia cynthia moth larvae,218 but does

not inhibit neuromuscular transmission on locust skeletal mus-

cle.219 The venom causes flaccid paralysis to ensue only after a

matter of minutes, maximizing at about 20 h in Galleria mello-

nella, while heart and gut muscles continue to function.220,221

Two protein toxins, Brh-I and Brh-V (approximate molecular

mass 73 kDa), were isolated from this venom and showed in-

secticidal activity when injected into six species of lepidopterous

larvae.222 The paralyzing components of B. hebetor venom are

thought to act by presynaptic blockage of the excitatory gluta-

matergic transmission at neuromuscular junctions, possibly by

inhibiting the release of synaptic vesicles.218,219

OTHER TARGETS FOR INSECTICIDALPEPTIDESThis section will deal with peptides with insecticidal activity

identified from arthropod venoms that act on molecular targets

other than those mentioned above, such as neurokinins. In

addition, it will be presented some insecticidal peptides whose

mechanism of action has not been fully understood so far.

Many neuroactive peptides with chemical structures and

physiological activities similar to bradykinin have been

reported in the venoms of wasps since the 1950’s.8,223,224 The

most active kinin isolated so far is Threonine6-Bradykinin

(Thr6-BK), which was previously found in venoms of social

wasp.22,225 Then, it was also identified in the venom of soli-

tary wasps Megascolia flavifrons226 and Colpa interrupta,

where this peptide is considered the most important toxic

component.227

The neurokinin Thr6-BK is a linear peptide with action

on CNS of cockroach.228,229 This kinin causes irreversible pa-

ralysis in insects through the presynaptic blockade of cholin-

ergic transmission caused by the non-competitive inhibition

of choline uptake.229,230 In this case, the blockade was attrib-

uted to a depletion of the readily releasable store of acetyl-

choline, similar to the activity of hemicholinium-3, a classic

choline uptake inhibitor.231,232 The irreversible effect of the

Thr6-BK in insects may be of interest for the use of this kinin

as a lead to new insecticides.230

Several bradykinin-like peptides that act on insect CNS

were also identified from the solitary wasp venom Paraves-

pula maculifrons and named vespulakinins.233 Modified ana-

logues of vespulakinin 1 and 2 provoked irreversible blockade

of synaptic transmission in cockroaches. Probably, these ana-

logues act similarly to Thr6-BK and BK by the blockage of

the excitatory nicotinic transmission with a concurrent acti-

vation of the inhibitory (GABA-ergic) system.231

The LaIT1 is a short chain toxin, with 36 amino acid resi-

dues and mass weight of 4200 Da, isolated from the scorpion







Liocheles australasiae (Hemiscorpiidae).234 This toxin fea-

tures two intramolecular disulfide bonds and presents two-

stranded antiparallel b-sheet and an ICK-like fold (Figure

1D, PDB code 2LDS).235 This peptide is toxic to crickets and

not toxic against mice (1 lg by i.c.v. injection), which sug-

gests its selectivity to insects. LaIT1 is similar to the putative

peptide OcyC10236 (64% identical) identified from the Bra-

zilian scorpion Opisthacanthus cayaporum, and 91% with

U1-liotoxin-Lw1a,90 from Liocheles waigiensis. The U1-lio-

toxin-Lw1a has no effects on Na+, K+, and Ca2+ currents

when tested on cockroach DUM neurons. The target of these

toxins is still unknown, but they seem to have activity only

against insects and are harmless to mammals. It is possible

that the peptides similar to LaIT1 target potassium or cal-

cium channels on insect, as shown for some spider toxins,

such as J-ACTX-Hv1c.235

The huwentoxin II (HWTX-II or U1-TRTX-Hh1a), iso-

lated from the venom of the spider Haplopelma schmidti, is a

37 amino acid residue long peptide with three disulfide

bonds.237 This toxin presents low insecticidal activity (ED50

of 127 6 54 lg g�1 in cockroaches), causing a dose-depend-

ent paralysis that lasts about 6 h. It also blocks neuromuscu-

lar transmission in isolated mouse phrenic nerve diaphragm

preparations.237 Differently from most of spider insecticidal

neurotoxins elucidated thus far, HWTX-II adopts a novel

scaffold, which conforms to the disulfide-directed b-hairpinmotif, without the typical cystine knot (Figure 1D, PDB code

1I25). It contains two b-turns and a double-stranded anti-

parallel b-sheet cross-linked by two disulfide bonds.129,238,239

With sequence similarity to HWTX-II, Ba1 (U1-TRTX-Ba1a)

and Ba2 (U1-TRTX-Ba1b), with 39 amino acid residues and

three disulfide bonds, from the spider Brachypelma ruhnaui

(¼ B. albiceps), are also toxic to insects. Ba1 and Ba2 were

not toxic to mice (3 and 20 lg/mouse, by intracranial and in-

traperitoneal route, respectively) and did not modify currents

in insect and vertebrate cloned Nav channels.240

APPLICATION IN AGRIBUSINESSThe increasing discovery of new insecticidal toxins and the

study of the interaction of these compounds with different

molecular targets have provided a promising potential for

the scientific community and agricultural industries. Due to

the excessive use of chemical insecticides, many insects have

developed resistance to insecticides with action on Nav chan-

nels, such as pyrethroid and other chemical insecticides (see

reviews241,242). In addition, mutations that confer resistance

to these compounds, named knockdown resistance (kdr and

super-kdr) are more sensitive to certain chemical insecticides

and peptide toxins that act on remote sites of the chan-

nel.67,243,244 Toxins that act on other molecular targets have

also offered a high potential for the design of new biopesti-

cides.

Pest control techniques with less environmental impact

have been sought to keep the pest species below the eco-

nomic-damage threshold, while leaving the other fauna spe-

cies undisturbed.245 These techniques include mechanical

methods, such as barriers or traps to capture the pest insects,

and biological methods, among which stand out the repro-

duction of natural predators of the host insects, the use of

biopesticides, such as Bacillus thuringiensis or entomopatho-

genic fungi, or the interruption of the reproductive process

of the pest through the insertion of sterile individuals.246–250

Another strategy that has been successfully field-tested is

the use of specific insect viruses that are designed to encode

insect-selective neurotoxins.251 Many studies regarding this

approach have used the baculoviruses, which are character-

ized by a large circular double-stranded DNA and enveloped

rod-shaped virions that are pathogenic to arthropods, mainly

holometabolous insects, such as Lepidoptera, and that do

not infect plants or vertebrates.252,253 Due to their high speci-

ficity, the baculoviruses have been considered safe agents in

the biological control of host insects, as they do not affect the

nontargeted populations, differently from most chemical

pesticides,222 which makes them ideal for incorporation into

integrated pest management programs.245 Two genera, the

Nucleopolyhedrovirus (NPV) and Granulovirus, make up the

Baculoviridae family. NPVs are arthropod specific viruses

that have long been used as natural insect biological control

agents for protection of numerous crop plants.

The most widely studied baculovirus is the Autographa

californica multiple nucleopolyhedrovirus (AcMNPV).

Among different NPVs, AcMNPV is known to infect *39

species of Lepidoptera including important pests in the gen-

era Heliothis (the tobacco plant pest), Spodoptera (feeds on

grasses and small grain crops), and Trichoplusia (feeds on

crucifers). Since the 80s, the AcMNPVs have been extensively

used in Brazil for the control of the velvet bean caterpillar

Anticarsia gemmatalis, which causes serious damage to soy-

bean crops in Latin America and southeastern United

States.254,255 The success of this project in Brazil revived the

interest in baculovirus as a biopesticide and gradually many

countries have begun to increase the area of fields and forests

to be experimentally protected by baculovirus pesti-

cides.256,257

However, the biggest limitations of baculovirus are the

lack of field persistence and the long latency time to exert its

insecticidal activity when compared to classical chemical

insecticides. The wild-type NPVs usually take from days to

weeks to kill their targets from the point of infection and

398 Schwartz et al.


during this time, the insect continues to feed, damaging the

plantations.252 This gap has been addressed by molecular en-

gineering techniques through the heterologous expression of

insect-selective neurotoxins from venomous animals, such as

scorpions,251,258–261 spiders,262–264 and sea anemones.262,263

All of them proved to reduce the time interval between appli-

cation of the virus and the death of the insect. The insertion

of the gene encoding for the toxin does not appear to alter

the virus specificity or its ability to replicate.

Previous studies with recombinant AcMNPVs expressing

the chimeric gene that encodes the l-agatoxin-IV (l-AGTX-Aa1d), from the spider A. aperta, with a melittin signal

sequence from the honey bee Apis mellifera, showed that lar-

vae infected with the genetically enhanced viruses died more

quickly than those infected with wild-type viruses.263 More-

over, a reduction in the median time to cessation of feeding

(FT50) was observed for the recombinant AcMNPV express-

ing both the toxins l-Aga-IV and As II - the latter from the

sea anemone Anemonia sulcata - compared with the single

toxin expressing viruses, thus having some advantage in

expressing two synergistic toxins simultaneously.262 A

recombinant AcMNPV expressing two scorpion toxins, both

excitatory LqhIT1 and depressant LqhIT2, also resulted in

synergistic effects with marked reduction in ET50 (mean time

to paralyze effectively or kill 50% of the test larvae).265

Other two recombinant AcMNPVs expressing insect-spe-

cific neurotoxin genes from the spiders Tegenaria agrestis and

Diguetia canities significantly reduced both the FT50 and the

median survival time (ST50) in the three infected lepidop-

teran pests Trichoplusiani (Hubner), Spodoptera exigua

(Hubner), and Heliothis virescens (Fabricius) when compared

with those larvae infected by wild-type AcMNPVs.266

A synthetic gene containing a lepidopteran-selective toxin,

Mesobuthus tamulus insect-selective toxin (ButaIT), in frame

to the bombyxin signal sequence, was engineered into a poly-

hedrin positive AcMNPV under the control of the p10 pro-

moter. This recombinant NPV (ButaIT-NPV) showed

enhanced insecticidal activity on the larvae of Heliothis vires-

cens as demonstrated by a significant reduction in the ST50

and a greater reduction in feeding damage as compared with

the wild-type AcMNPV.267

ButaIT was fused N-terminally to a snowdrop lectin (Gal-

anthus nivalis agglutinin; GNA) polypeptide and expressed

in the yeast Pichia pastoris. The recombinant ButaIT/GNA

was tested on larvae of the tomato moth (Lacanobia oleracea)

and proved acutely toxic when injected into the larvae—

causing slow paralysis, and leading to mortality or decreased

growth – and chronically toxic when fed to the larvae—caus-

ing decreased survival and weight gain under conditions

where GNA alone was effectively nontoxic.268

Poneratoxin, from Paraponera clavata, appears to be a

good candidate for the construction of an insecticidal bacu-

lovirus able to immobilize the infected insect. The baculovi-

rus engineered to express poneratoxin reduced the survival

average time of S. frugiperda larvae by 25 h compared with

unmodified virus.44

Khan et al.269 showed that the toxin x-ACTX-Hv1a, from

the spider Hadronyche versuta,172 retains its biological activ-

ity antagonist of insect Cav channel when expressed in a het-

erologous system. Expressed as a fusion protein in E. coli, the

purified toxin fusion immobilized and killed the larvae of

two recalcitrant agricultural pests - Helicover paarmigera

(cotton bollworm) and Spodoptera littoralis (Egyptian army-

worm) caterpillars - when applied topically. When expressed

heterologously in tobacco (Nicotiana tabacum) plants, the

toxin also retained its neurological toxicity in at least these

two insect species, inducing lack of coordination, disorienta-

tion, and uncontrolled movements. Typically, the larvae died

within 48 h after starting feeding on the genetically modified

leaves. There was no apparent deleterious effect of transgene

expression on plant growth, development, and fertility.269

Hernandez-Campuzano et al.270 conducted a similar

study, cloning and expressing the toxin Magi-6 from the

venom of the spider Macrothele gigas in tobacco plants. No

morphological alterations in the different transgenic lines

were observed nor there was no change in plant growth. The

transgenic lines were significantly more resistant to the her-

bivorous insect larvae Spodoptera frugiperda than the wild-

type plants. The symptoms of mortality consisted in cessa-

tion of feeding and uncontrolled movements, followed by pa-

ralysis. Since the oral administration of active insect neuro-

peptides is unlikely to be successful as the insect gut enzymes

would probably degrade them, this study suggests the possi-

ble action of plant lectins as a carrier to deliver peptides to

the hemolymph of the target insects. The mode of action of

this toxin is still unknown and due to its activity on mice,20

an initial application of Magi-6 is proposed in inedible crops,

such as the cotton of ornamental plants, although toxicity

tests on mammals are still required.270

Many studies aiming at the use of insecticide toxins asso-

ciated with baculovirus have been carried out in the 90s. De-

spite of that, the new studies presented in this section and

the existence of records in recent worldwide patent databases

(U.S. Pat. No. 7,951,929 and U.S. Pat. No. 7,880,060) show

that this may be a promising field for future research and de-

velopment of new compounds with potential use in agricul-

ture.256,265 In addition, the use of toxins associated with the







baculovirus can expand beyond the crops, since there are

baculovirus that target vectors of human diseases, including

the NPVs specific to mosquitoes of the genera Aedes, Anophe-

les, and Culex.271–273

Regarding the biosafety of recombinant baculoviruses,

there is no evidence that they represent greater threats to the

environment than the wild-type baculovirus. Furthermore,

biopesticides based on baculovirus formulations represent a

much lower risk to the environment than conventional

chemical pesticides (for review255,256). Nevertheless, both

recombinant baculovirus and plant-engineered systems

require special attention in their production and application.

CONCLUSIONThe arthropod venoms possess a variety of neuroactive pep-

tides with high selectivity and specificity for insects. These

peptides may be involved in the control of almost all key

functions in insects and thus are prime targets for the devel-

opment of a novel generation of selective insecticides. With a

modest estimate of approximately 100 peptides per venom of

spiders and scorpions, and about 30 peptides per wasp

venom, and based on the extraordinary taxonomic diversity

of arthropods, including those poorly explored (centipedes

and ants, for example), a collection of millions of molecules

is to be expected, revealing the enormous potential of these

animals in the discovery of new insecticide compounds. A

large number of putative insecticidal peptides deposited in

public databases do not have its molecular target character-

ized yet.

A premise for the studies with heterologous systems to

become commercially applied is the increased disclosure

about the risks and benefits of chemical and biological pesti-

cides for the public. Moreover, some attitude change of law-

makers in some countries is also essential for the recombi-

nant biopesticides to gradually increase their share in pesti-

cide market.255,256

REFERENCES1. Jackson, H.; Usherwood, P. N. Trends Neurosci 1988, 11, 278–

283.

2. Nakajima, T. Regul Pept 1993, 46, 89–92.

3. Rash, L. D.; Hodgson, W. C. Toxicon 2002, 40, 225–254.

4. Jackson, H.; Parks, T. N. Annu Rev Neurosci 1989, 12, 405–414.

5. Pimenta, A. M.; De Lima, M. E. J Pept Sci 2005, 11, 670–676.

6. Usherwood, P. N.; Blagbrough, I. S. Pharmacol Ther 1991, 52,

245–268.

7. Beleboni, R. O.; Pizzo, A. B.; Fontana, A. C.; Carolino, R. O.

G.; Coutinho-Netto, J.; Santos, W. F. Eur J Pharmacol 2004,

493, 1–17.

8. Mortari, M. R.; Cunha, A. O.; Ferreira, L. B.; dos Santos, W. F.

Pharmacol Ther 2007, 114, 171–183.

9. Quistad, G. B.; Reuter, C. C.; Skinner, W. S.; Dennis, P. A.;

Suwanrumpha, S.; Fu, E. W. Toxicon 1991, 29, 329–336.

10. Bloomquist, J. R. Annu Rev Entomol 1996, 41, 163–190.

11. Harvey, A. L.; Bradley, K. N.; Cochran, S. A.; Rowan, E. G.;

Pratt, J. A.; Quillfeldt, J. A.; Jerusalinsky, D. A. Toxicon 1998,

36, 1635–1640.

12. Estrada, G.; Villegas, E.; Corzo, G. Nat Prod Rep 2007, 24,

145–161.

13. Corzo, G.; Villegas, E.; Gomez-Lagunas, F.; Possani, L. D.;

Belokoneva, O. S.; Nakajima, T. J Biol Chem 2002, 277,

23627–23637.

14. Herzig, V.; Wood, D. L.; Newell, F.; Chaumeil, P. A.; Kaas, Q.;

Binford, G. J.; Nicholson, G. M.; Gorse, D.; King, G. F. Nucleic

Acids Res 2011, 39, D653–657.

15. Tytgat, J.; Chandy, K. G.; Garcia, M. L.; Gutman, G. A.; Mar-

tin-Eauclaire, M. F.; van der Walt, J. J.; Possani, L. D. Trends

Pharmacol Sci 1999, 20, 444–447.

16. Rodriguez de la Vega, R. C.; Possani, L. D. Toxicon 2005, 46,

831–844.

17. Nicholson, G. M. Toxicon 2007, 49, 490–512.

18. Gurevitz, M.; Karbat, I.; Cohen, L.; Ilan, N.; Kahn, R.; Turkov,

M.; Stankiewicz, M.; Stuhmer, W.; Dong, K.; Gordon, D. Toxi-

con 2007, 49, 473–489.

19. De Lima, M. E.; Figueiredo, S. G.; Pimenta, A. M.; Santos, D.

M.; Borges, M. H.; Cordeiro, M. N.; Richardson, M.; Oliveira,

L. C.; Stankiewicz, M.; Pelhate, M. Comp Biochem Physiol C

2007, 146, 264–279.

20. Corzo, G.; Escoubas, P. Cell Mol Life Sci 2003, 60, 2409–2426.

21. Meinwald, J.; Eisner, T. Proc Natl Acad Sci USA 1995, 92, 14–

18.

22. Nakajima, T.; Uzu, S.; Wakamatsu, K.; Saito, K.; Miyazawa, T.;

Yasuhara, T.; Tsukamoto, Y.; Fujino, M. Biopolymers 1986,

25Suppl, S115–121.

23. Cascone, O.; Amaral, V.; Ferrara, P.; Vita, N.; Guillemot, J. C.;

Diaz, L. E. Toxicon 1995, 33, 659–665.

24. Hoffman, D. R. J Allergy Clin Immunol 1993, 92, 707–716.

25. Pirpignani, M. L.; Rivera, E.; Hellman, U.; Biscoglio de Jime-

nez Bonino, M. Arch Biochem Biophys 2002, 407, 224–230.

26. Kolarich, D.; Leonard, R.; Hemmer, W.; Altmann, F. FEBS J

2005, 272, 5182–5190.

27. Soldatova, L.; Kochoumian, L.; King, T. P. FEBS Lett 1993,

320, 145–149.

28. Abe, T.; Sugita, M.; Fujikura, T.; Hiyoshi, J.; Akasu, M. Toxi-

con 2000, 38, 1803–1816.

29. Costa, H.; Palma, M. S. Toxicon 2000, 38, 1367–1379.

30. Brigatte, P.; Cury, Y.; de Souza, B. M.; Baptista-Saidemberg, N.

B.; Saidemberg, D. M.; Gutierrez, V. P.; Palma, M. S. Amino

Acids 2011, 40, 101–111.

31. Baptista-Saidemberg, N. B.; Saidemberg, D. M.; Palma, M. S. J

Proteomics 2011, 74, 2123–2137.

32. Souza, B. M.; Marques, M. R.; Tomazela, D. M.; Eberlin, M.

N.; Mendes, M. A.; Palma, M. S. Rapid Commun Mass Spec-

trom 2004, 18, 1095–1102.

33. Mendes, M. A.; de Souza, B. M.; Marques, M. R.; Palma, M. S.

Toxicon 2004, 44, 67–74.

34. Habermann, E. Pharmacol Ther 1984, 25, 255–270.

35. Son, D. J.; Kang, J.; Kim, T. J.; Song, H. S.; Sung, K. J.; Yun do,

Y.; Hong, J. T. J Toxicol Environ Health A 2007, 70, 1350–

400 Schwartz et al.


1355.

36. Orivel, J.; Redeker, V.; Le Caer, J. P.; Krier, F.; Revol-Junelles, A.

M.; Longeon, A.; Chaffotte, A.; Dejean, A.; Rossier, J. J Biol

Chem 2001, 276, 17823–17829.

37. Piek, T.; Duval, A.; Hue, B.; Karst, H.; Lapied, B.; Mantel, P.;

Nakajima, T.; Pelhate, M.; Schmidt, J. O. Comp Biochem

Physiol C 1991, 99, 487–495.

38. Mohamed, A. H.; Abu-Sinna, G.; El-Shabaka, H. A.; El-Aal, A.

A. Toxicon 1983, 21, 371–377.

39. Gutierrez Mdel, C.; Abarca, C.; Possani, L. D. Comp Biochem

Physiol C 2003, 135, 205–214.

40. Malta, M. B.; Lira, M. S.; Soares, S. L.; Rocha, G. C.; Knysak,

I.; Martins, R.; Guizze, S. P.; Santoro, M. L.; Barbaro, K. C.

Toxicon 2008, 52, 255–263.

41. Stankiewicz, M.; Hamon, A.; Benkhalifa, R.; Kadziela, W.;

Hue, B.; Lucas, S.; Mebs, D.; Pelhate, M. Toxicon 1999, 37,

1431–1445.

42. Rates, B.; Bemquerer, M. P.; Richardson, M.; Borges, M. H.;

Morales, R. A.; De Lima, M. E.; Pimenta, A. M. Toxicon 2007,

49, 810–826.

43. King, G. F. Toxicon 2007, 49, 513–530.

44. Szolajska, E.; Poznanski, J.; Ferber, M. L.; Michalik, J.; Gout,

E.; Fender, P.; Bailly, I.; Dublet, B.; Chroboczek, J. Eur J

Biochem 2004, 271, 2127–2136.

45. Stevens, M.; Peigneur, S.; Tytgat, J. Front Pharmacol 2011, 2,

71.

46. Chahine, M.; Chen, L. Q.; Fotouhi, N.; Walsky, R.; Fry, D.;

Santarelli, V.; Horn, R.; Kallen, R. G. Receptors Channels

1995, 3, 161–174.

47. Chahine, M.; Sirois, J.; Marcotte, P.; Chen, L.; Kallen, R. G.

Biophys J 1998, 75, 236–246.

48. Narahashi, T.; Moore, J. W.; Scott, W. R. J Gen Physiol 1964,

47, 965–974.

49. Ritchie, J. M.; Rogart, R. B. Rev Physiol Biochem Pharmacol

1977, 79, 1–50.

50. Terlau, H.; Heinemann, S. H.; Stuhmer, W.; Pusch, M.; Conti,

F.; Imoto, K.; Numa, S. FEBS Lett 1991, 293, 93–96.

51. Mattei, C.; Molgo, J.; Legrand, A. M.; Benoit, E. J De La Soc

De Biol 1999, 193, 329–344.

52. Meunier, F. A.; Mattei, C.; Molgo, J. Prog Mol Subcell Biol

2009, 46, 159–186.

53. Rogers, J. C.; Qu, Y.; Tanada, T. N.; Scheuer, T.; Catterall, W.

A. J Biol Chem 1996, 271, 15950–15962.

54. Sheets, M. F.; Kyle, J. W.; Kallen, R. G.; Hanck, D. A. Biophys J

1999, 77, 747–757.

55. Corzo, G.; Gilles, N.; Satake, H.; Villegas, E.; Dai, L.; Naka-

jima, T.; Haupt, J. FEBS Lett 2003, 547, 43–50.

56. De Lima, M. E.; Stankiewicz, M.; Hamon, A.; de Figueiredo, S.

G.; Cordeiro, M. N.; Diniz, C. R.; Martin-Eauclaire, M.; Pel-

hate, M. J Insect Physiol 2002, 48, 53–61.

57. Liu, Z. R.; Ye, P.; Ji, Y. H. Protein Cell 2011, 2, 437–444.

58. Billen, B.; Tytgat, J. In Animal Toxins: State of the Art-Perspec-

tives on Health and Biotechnology; Lima, M. E., Ed.; Belo

Horizonte, Brasil, 2009, p 137–166.

59. Leipold, E.; Hansel, A.; Olivera, B. M.; Terlau, H.; Heinemann,

S. H. FEBS Lett 2005, 579, 3881–3884.

60. Bloomquist, J. R. Comp Biochem Physiol C 1993, 106, 301–

314.

61. Salkoff, L.; Butler, A.; Wei, A.; Scavarda, N.; Giffen, K.; Ifune,

C.; Goodman, R.; Mandel, G. Science 1987, 237, 744–749.

62. Loughney, K.; Kreber, R.; Ganetzky, B. Cell 1989, 58, 1143–

1154.

63. Zhang, T.; Liu, Z.; Song, W.; Du, Y.; Dong, K. Insect Biochem

Mol Biol 2011, 41, 451–458.

64. Feng, G.; Deak, P.; Chopra, M.; Hall, L. M. Cell 1995, 82,

1001–1011.

65. Warmke, J. W.; Reenan, R. A.; Wang, P.; Qian, S.; Arena, J. P.;

Wang, J.; Wunderler, D.; Liu, K.; Kaczorowski, G. J.; Van der

Ploeg, L. H.; Ganetzky, B.; Cohen, C. J. J Gen Physiol 1997,

110, 119–133.

66. Dong, K. Invert Neurosci 2007, 7, 17–30.

67. Zlotkin, E. Annu Rev Entomol 1999, 44, 429–455.

68. Li, D.; Xiao, Y.; Hu, W.; Xie, J.; Bosmans, F.; Tytgat, J.; Liang,

S. FEBS Lett 2003, 555, 616–622.

69. Figueiredo, S. G.; Garcia, M. E.; Valentim, A. C.; Cordeiro, M.

N.; Diniz, C. R.; Richardson, M. Toxicon 1995, 33, 83–93.

70. Penaforte, C. L.; Prado, V. F.; Prado, M. A.; Romano-Silva, M.

A.; Guimaraes, P. E.; De Marco, L.; Gomez, M. V.; Kalapotha-

kis, E. Toxicon 2000, 38, 1443–1449.

71. Sheumack, D. D.; Claassens, R.; Whiteley, N. M.; Howden, M.

E. FEBS Lett 1985, 181, 154–156.

72. Brown, M. R.; Sheumack, D. D.; Tyler, M. I.; Howden, M. E.

Biochem J 1988, 250, 401–405.

73. Grolleau, F.; Stankiewicz, M.; Birinyi-Strachan, L.; Wang, X.

H.; Nicholson, G. M.; Pelhate, M.; Lapied, B. J Exp Biol 2001,

204, 711–721.

74. Little, M. J.; Wilson, H.; Zappia, C.; Cestele, S.; Tyler, M. I.;

Martin-Eauclaire, M. F.; Gordon, D.; Nicholson, G. M. FEBS

Lett 1998, 439, 246–252.

75. Nicholson, G. M.; Little, M. J.; Tyler, M.; Narahashi, T. Toxi-

con 1996, 34, 1443–1453.

76. Nicholson, G. M.; Little, M. J.; Birinyi-Strachan, L. C. Toxicon

2004, 43, 587–599.

77. Nicholson, G. M.; Walsh, R.; Little, M. J.; Tyler, M. I. Pflugers

Arch 1998, 436, 117–126.

78. Nicholson, G. M.; Willow, M.; Howden, M. E.; Narahashi, T.

Pflugers Arch 1994, 428, 400–409.

79. Zeng, X. Z.; Zhu, Q.; Liang, S. P. Acta Biochim Biophys Sinica

2005, 37, 567–572.

80. Xiao, Y.; Li, J.; Deng, M.; Dai, C.; Liang, S. Toxicon 2007, 50,

507–517.

81. Corzo, G.; Escoubas, P.; Stankiewicz, M.; Pelhate, M.; Kristen-

sen, C. P.; Nakajima, T. Eur J Biochem 2000, 267, 5783–5795.

82. Corzo, G.; Escoubas, P.; Villegas, E.; Karbat, I.; Gordon, D.;

Gurevitz, M.; Nakajima, T.; Gilles, N. Biochemistry 2005, 44,

1542–1549.

83. Ferrat, G.; Bosmans, F.; Tytgat, J.; Pimentel, C.; Chagot, B.;

Gilles, N.; Nakajima, T.; Darbon, H.; Corzo, G. Proteins 2005,

59, 368–379.

84. Skinner, W. S.; Adams, M. E.; Quistad, G. B.; Kataoka, H.;

Cesarin, B. J.; Enderlin, F. E.; Schooley, D. A. J Biol Chem

1989, 264, 2150–2155.

85. Adams, M. E.; Herold, E. E.; Venema, V. J. J Comp Physiol [A]

1989, 164, 333–342.







86. Stapleton, A.; Blankenship, D. T.; Ackermann, B. L.; Chen, T.

M.; Gorder, G. W.; Manley, G. D.; Palfreyman, M. G.; Coutant,

J. E.; Cardin, A. D. J Biol Chem 1990, 265, 2054–2059.

87. Fletcher, J. I.; Chapman, B. E.; Mackay, J. P.; Howden, M. E.;

King, G. F. Structure 1997, 5, 1525–1535.

88. Omecinsky, D. O.; Holub, K. E.; Adams, M. E.; Reily, M. D.

Biochemistry 1996, 35, 2836–2844.

89. Gilles, N.; Harrison, G.; Karbat, I.; Gurevitz, M.; Nicholson, G.

M.; Gordon, D. Eur J Biochem 2002, 269, 1500–1510.

90. Smith, J. J.; Hill, J. M.; Little, M. J.; Nicholson, G. M.; King, G.

F.; Alewood, P. F. Proc Natl Acad Sci USA 2011, 108, 10478–

10483.

91. Xiao, Y.; Tang, J.; Hu, W.; Xie, J.; Maertens, C.; Tytgat, J.;

Liang, S. J Biol Chem 2005, 280, 12069–12076.

92. Yuan, C.; Yang, S.; Liao, Z.; Liang, S. Biochem Biophys Res

Commun 2007, 352, 799–804.

93. Cestele, S.; Catterall, W. A. Biochimie 2000, 82, 883–892.

94. Possani, L. D.; Merino, E.; Corona, M.; Bolivar, F.; Becerril, B.

Biochimie 2000, 82, 861–868.

95. Legros, C.; Ceard, B.; Vacher, H.; Marchot, P.; Bougis, P. E.; Mar-

tin-Eauclaire, M. F. Biochim Biophys Acta 2005, 1723, 91–99.

96. Sautiere, P.; Cestele, S.; Kopeyan, C.; Martinage, A.; Drobecq,

H.; Doljansky, Y.; Gordon, D. Toxicon 1998, 36, 1141–1154.

97. Eitan, M.; Fowler, E.; Herrmann, R.; Duval, A.; Pelhate, M.;

Zlotkin, E. Biochemistry 1990, 29, 5941–5947.

98. Gordon, D.; Martin-Eauclaire, M. F.; Cestele, S.; Kopeyan, C.;

Carlier, E.; Khalifa, R. B.; Pelhate, M.; Rochat, H. J Biol Chem

1996, 271, 8034–8045.

99. Krimm, I.; Gilles, N.; Sautiere, P.; Stankiewicz, M.; Pelhate,

M.; Gordon, D.; Lancelin, J. M. J Mol Biol 1999, 285, 1749–

1763.

100. Tugarinov, V.; Kustanovich, I.; Zilberberg, N.; Gurevitz, M.;

Anglister, J. Biochemistry 1997, 36, 2414–2424.

101. Zilberberg, N.; Gordon, D.; Pelhate, M.; Adams, M. E.; Norris,

T. M.; Zlotkin, E.; Gurevitz, M. Biochemistry 1996, 35, 10215–

10222.

102. Borchani, L.; Stankiewicz, M.; Kopeyan, C.; Mansuelle, P.;

Kharrat, R.; Cestele, S.; Karoui, H.; Rochat, H.; Pelhate, M.; el

Ayeb, M. Toxicon 1997, 35, 365–382.

103. Arnon, T.; Potikha, T.; Sher, D.; Elazar, M.; Mao, W.; Tal, T.;

Bosmans, F.; Tytgat, J.; Ben-Arie, N.; Zlotkin, E. Insect Bio-

chem Mol Biol 2005, 35, 187–195.

104. Gordon, D.; Ilan, N.; Zilberberg, N.; Gilles, N.; Urbach, D.;

Cohen, L.; Karbat, I.; Froy, O.; Gaathon, A.; Kallen, R. G.; Ben-

veniste, M.; Gurevitz, M. Eur J Biochem 2003, 270, 2663–

2670.

105. Rodriguez de la Vega, R. C.; Possani, L. D. Toxicon 2007, 49,

171–180.

106. Vazquez, A.; Tapia, J. V.; Eliason, W. K.; Martin, B. M.; Lebre-

ton, F.; Delepierre, M.; Possani, L. D.; Becerril, B. Toxicon

1995, 33, 1161–1170.

107. Pintar, A.; Possani, L. D.; Delepierre, M. J Mol Biol 1999, 287,

359–367.

108. Martin, M. F.; Garcia y Perez, L. G.; el Ayeb, M.; Kopeyan, C.;

Bechis, G.; Jover, E.; Rochat, H. J Biol Chem 1987, 262, 4452–

4459.

109. Possani, L. D.; Becerril, B.; Delepierre, M.; Tytgat, J. Eur J Bio-

chem 1999, 264, 287–300.

110. Froy, O.; Zilberberg, N.; Gordon, D.; Turkov, M.; Gilles, N.;

Stankiewicz, M.; Pelhate, M.; Loret, E.; Oren, D. A.; Shaanan,

B.; Gurevitz, M. J Biol Chem 1999, 274, 5769–5776.

111. Loret, E. P.; Martin-Eauclaire, M. F.; Mansuelle, P.; Sampieri,

F.; Granier, C.; Rochat, H. Biochemistry 1991, 30, 633–640.

112. Ji, Y. H.; Mansuelle, P.; Xu, K.; Granier, C.; Kopeyan, C.; Tera-

kawa, S.; Rochat, H. Sci China Ser B Chem Life Sci Earth Sci

1994, 37, 42–49.

113. Tong, X.; Yao, J.; He, F.; Chen, X.; Zheng, X.; Xie, C.; Wu, G.;

Zhang, N.; Ding, J.; Wu, H. Biochem Biophys Res Commun

2006, 349, 890–899.

114. Zlotkin, E.; Eitan, M.; Bindokas, V. P.; Adams, M. E.; Moyer,

M.; Burkhart, W.; Fowler, E. Biochemistry 1991, 30, 4814–

4821.

115. Ji, Y. H.; Hattori, H.; Xu, K.; Terakawa, S. Sci China Ser B

Chem Life Sci Earth Sci 1994, 37, 955–963.

116. Ji, Y. H.; Terakawa, S.; Xu, K. Chin Sci Bull 1994, 39, 945–949.

117. Li, Y. J.; Tan, Z. Y.; Ji, Y. H. Neurosci Res 2000, 38, 257–264.

118. He, H.; Liu, Z.; Dong, B.; Zhang, J.; Shu, X.; Zhou, J.; Ji, Y.

PloS ONE 2011, 6, e14510.

119. Guerrero-Vargas, J. A.; Mourao, C. B.; Quintero-Hernandez,

V.; Possani, L. D.; Schwartz, E. F. PloS ONE 2012, 7, e30478.

120. Schiavon, E.; Stevens, M.; Zaharenko, A. J.; Konno, K.; Tytgat,

J.; Wanke, E. FEBS J 2010, 277, 918–930.

121. Ishaaya, I.; Kontsedalov, S.; Horowitz, A. R. Arch Insect Bio-

chem Physiol 2005, 58, 192–199.

122. Dhadialla, T. S.; Carlson, G. R.; Le, D. P. Annu Rev Entomol

1998, 43, 545–569.

123. Salgado, V. L. Pestic Biochem Physiol 1992, 43, 1–13.

124. Judge, S. I.; Smith, P. J.; Stewart, P. E.; Bever, C. T., Jr. Recent

Patents CNS Drug Discov 2007, 2, 200–228.

125. Vergara, C.; Latorre, R.; Marrion, N. V.; Adelman, J. P. Curr

Opin Neurobiol 1998, 8, 321–329.

126. Toro, L.; Wallner, M.; Meera, P.; Tanaka, Y. News Physiol Sci

1998, 13, 112–117.

127. Ghatta, S.; Nimmagadda, D.; Xu, X.; O’Rourke, S. T. Pharma-

col Ther 2006, 110, 103–116.

128. Gunning, S. J.; Maggio, F.; Windley, M. J.; Valenzuela, S. M.;

King, G. F.; Nicholson, G. M. FEBS J 2008, 275, 4045–4059.

129. Wang, X.; Connor, M.; Smith, R.; Maciejewski, M. W.; How-

den, M. E.; Nicholson, G. M.; Christie, M. J.; King, G. F. Nat

Struct Biol 2000, 7, 505–513.

130. Maggio, F.; King, G. F. Toxicon 2002, 40, 1355–1361.

131. Maggio, F.; King, G. F. J Biol Chem 2002, 277, 22806–22813.

132. Tedford, H. W.; Maggio, F.; Reenan, R. A.; King, G. Peptides

2007, 28, 51–56.

133. Windley, M. J.; Escoubas, P.; Valenzuela, S. M.; Nicholson, G.

M. Mol Pharmacol 2011, 80, 1–13.

134. Miller, C.; Moczydlowski, E.; Latorre, R.; Phillips, M. Nature

1985, 313, 316–318.

135. Lambert, P.; Kuroda, H.; Chino, N.; Watanabe, T. X.; Kimura,

T.; Sakakibara, S. Biochem Biophys Res Commun 1990, 170,

684–690.

136. Galvez, A.; Gimenez-Gallego, G.; Reuben, J. P.; Roy-Contan-

cin, L.; Feigenbaum, P.; Kaczorowski, G. J.; Garcia, M. L. J Biol

Chem 1990, 265, 11083–11090.

137. Giangiacomo, K. M.; Garcia, M. L.; McManus, O. B. Biochem-

istry 1992, 31, 6719–6727.

138. Derst, C.; Messutat, S.; Walther, C.; Eckert, M.; Heinemann, S.

H.; Wicher, D. Eur J Neurosci 2003, 17, 1197–1212.

402 Schwartz et al.


https://www.researchgate.net/publication/21125534_Functional_duality_and_structural_uniqueness_of_depressant_insect-selective_neurotoxins?el=1_x_8&enrichId=rgreq-5ae8bb89-6f3c-44c3-a86e-299b5b5dda46&enrichSource=Y292ZXJQYWdlOzIzMzc5OTM2NTtBUzoxMDI3NTYyODMwNjAyMjdAMTQwMTUxMDQxNTM0MA==



139. Xu, C. Q.; Brone, B.; Wicher, D.; Bozkurt, O.; Lu, W. Y.; Huys,

I.; Han, Y. H.; Tytgat, J.; Van Kerkhove, E.; Chi, C. W. J Biol

Chem 2004, 279, 34562–34569.

140. MacKinnon, R.; Reinhart, P. H.; White, M. M. Neuron 1988,

1, 997–1001.

141. Gao, B.; Peigneur, S.; Tytgat, J.; Zhu, S. Biochimie 2010, 92,

1847–1853.

142. Schwartz, E. F.; Schwartz, C. A.; Gomez-Lagunas, F.; Zamudio,

F. Z.; Possani, L. D. Toxicon 2006, 48, 1046–1053.

143. Batista, C. V.; D’Suze, G.; Gomez-Lagunas, F.; Zamudio, F. Z.;

Encarnacion, S.; Sevcik, C.; Possani, L. D. Proteomics 2006, 6,

3718–3727.

144. Diego-Garcia, E.; Batista, C. V.; Garcia-Gomez, B. I.; Lucas, S.;

Candido, D. M.; Gomez-Lagunas, F.; Possani, L. D. Toxicon

2005, 45, 273–283.

145. Gomez-Lagunas, F.; Olamendi-Portugal, T.; Possani, L. D.

FEBS Lett 1997, 400, 197–200.

146. Gomez-Lagunas, F.; Melishchuk, A.; Armstrong, C. M. Proc

Natl Acad Sci USA 2003, 100, 347–351.

147. Batista, C. V.; Gomez-Lagunas, F.; Lucas, S.; Possani, L. D.

FEBS Lett 2000, 486, 117–120.

148. Garcia, M. L.; Garcia-Calvo, M.; Hidalgo, P.; Lee, A.; MacKin-

non, R. Biochemistry 1994, 33, 6834–6839.

149. Garcia-Gomez, B. I.; Coronas, F. I.; Restano-Cassulini, R.;

Rodriguez, R. R.; Possani, L. D. Toxicon 2011, 58, 18–27.

150. Diego-Garcia, E.; Abdel-Mottaleb, Y.; Schwartz, E. F.; de la

Vega, R. C.; Tytgat, J.; Possani, L. D. Cell Mol Life Sci 2008, 65,

187–200.

151. Uawonggul, N.; Thammasirirak, S.; Chaveerach, A.;

Arkaravichien, T.; Bunyatratchata, W.; Ruangjirachuporn, W.;

Jearranaiprepame, P.; Nakamura, T.; Matsuda, M.;

Kobayashi, M.; Hattori, S.; Daduang, S. Toxicon 2007, 49, 19–

29.

152. Matsushita, N.; Miyashita, M.; Ichiki, Y.; Ogura, T.; Sakura-

dani, E.; Nakagawa, Y.; Shimizu, S.; Miyagawa, H. Biosci Bio-

technol Biochem 2009, 73, 2769–2772.

153. Diego-Garcia, E.; Schwartz, E. F.; D’Suze, G.; Gonzalez, S. A.;

Batista, C. V.; Garcia, B. I.; de la Vega, R. C.; Possani, L. D.

Peptides 2007, 28, 31–37.

154. Schwartz, E. F.; Diego-Garcia, E.; Rodriguez de la Vega, R. C.;

Possani, L. D. BMC Genomics 2007, 8, 119.

155. Gauldie, J.; Hanson, J. M.; Rumjanek, F. D.; Shipolini, R. A.;

Vernon, C. A. Eur J Biochem 1976, 61, 369–376.

156. Habermann, E. Science 1972, 177, 314–322.

157. Garcia, M. L.; Galvez, A.; Garcia-Calvo, M.; King, V. F.; Vaz-

quez, J.; Kaczorowski, G. J. J Bioenerg Biomembr 1991, 23,

615–646.

158. van der Staay, F. J.; Fanelli, R. J.; Blokland, A.; Schmidt, B. H.

Neurosci Biobehav Rev 1999, 23, 1087–1110.

159. Mpari, B.; Regaya, I.; Escoffier, G.; Mourre, C. J Integrative

Neurosci 2005, 4, 381–396.

160. Jin, W.; Lu, Z. Biochemistry 1998, 37, 13291–13299.

161. Berridge, M. J. J Physiol 1997, 499 (Pt 2), 291–306.

162. Bers, D. Excitation-Contraction Coupling & Cardiac Con-

tractile Force; Kluwer Academic Publishers: Amsterdam,

2001.

163. Sutko, J. L.; Airey, J. A.; Welch, W.; Ruest, L. Pharmacol Rev

1997, 49, 53–98.

164. Lahm, G. P.; Cordova, D.; Barry, J. D. Bioorg Med Chem 2009,

17, 4127–4133.

165. Sattelle, D. B.; Cordova, D.; Cheek, T. R. Invert Neurosci 2008,

8, 107–119.

166. Littleton, J. T.; Ganetzky, B. Neuron 2000, 26, 35–43.

167. Rieckhof, G. E.; Yoshihara, M.; Guan, Z.; Littleton, J. T. J Biol

Chem 2003, 278, 41099–41108.

168. Hoopengardner, B.; Bhalla, T.; Staber, C.; Reenan, R. Science

2003, 301, 832–836.

169. Smith, L. A.; Peixoto, A. A.; Hall, J. C. J Neurogenet 1998, 12,

227–240.

170. Eberl, D. F.; Ren, D.; Feng, G.; Lorenz, L. J.; Van Vactor, D.;

Hall, L. M. Genetics 1998, 148, 1159–1169.

171. Kawasaki, F.; Collins, S. C.; Ordway, R. W. J Neurosci 2002,

22, 5856–5864.

172. Fletcher, J. I.; Smith, R.; O’Donoghue, S. I.; Nilges, M.; Con-

nor, M.; Howden, M. E.; Christie, M. J.; King, G. F. Nat Struct

Biol 1997, 4, 559–566.

173. Tedford, H. W.; Fletcher, J. I.; King, G. F. J Biol Chem 2001,

276, 26568–26576.

174. Tedford, H. W.; Gilles, N.; Menez, A.; Doering, C. J.; Zamponi,

G. W.; King, G. F. J Biol Chem 2004, 279, 44133–44140.

175. Bloomquist, J. R. Invert Neurosci 2003, 5, 45–50.

176. Wang, X.; Smith, R.; Fletcher, J. I.; Wilson, H.; Wood, C. J.;

Howden, M. E.; King, G. F. Eur J Biochem 1999, 264, 488–

494.

177. Chong, Y.; Hayes, J. L.; Sollod, B.; Wen, S.; Wilson, D. T.;

Hains, P. G.; Hodgson, W. C.; Broady, K. W.; King, G. F.; Nich-

olson, G. M. Biochem Pharmacol 2007, 74, 623–638.

178. Wang, X. H.; Connor, M.; Wilson, D.; Wilson, H. I.; Nichol-

son, G. M.; Smith, R.; Shaw, D.; Mackay, J. P.; Alewood, P. F.;

Christie, M. J.; King, G. F. J Biol Chem 2001, 276, 40306–

40312.

179. Zhang, P. F.; Chen, P.; Hu, W. J.; Liang, S. P. Toxicon 2003, 42,

15–20.

180. Deng, M.; Luo, X.; Meng, E.; Xiao, Y.; Liang, S. Eur J Pharma-

col 2008, 582, 12–16.

181. Kuhn-Nentwig, L.; Schaller, J.; Nentwig, W. Toxicon 1994, 32,

287–302.

182. Kuhn-Nentwig, L.; Schaller, J.; Kampfer, U.; Imboden, H.;

Malli, H.; Nentwig, W. Arch Insect Biochem Physiol 2000, 44,

101–111.

183. Kubista, H.; Mafra, R. A.; Chong, Y.; Nicholson, G. M.; Beirao,

P. S.; Cruz, J. S.; Boehm, S.; Nentwig, W.; Kuhn-Nentwig, L.

Neuropharmacology 2007, 52, 1650–1662.

184. Adams, M. E. Toxicon 2004, 43, 509–525.

185. Branton, W. D.; Kolton, L.; Jan, Y. N.; Jan, L. Y. J Neurosci

1987, 7, 4195–4200.

186. Kuromi, H.; Honda, A.; Kidokoro, Y. Neuron 2004, 41, 101–

111.

187. Leung, H. T.; Branton, W. D.; Phillips, H. S.; Jan, L.; Byerly, L.

Neuron 1989, 3, 767–772.

188. Diao, J.; Lin, Y.; Tang, J.; Liang, S. Toxicon 2003, 42, 715–723.

189. Mintz, I. M.; Venema, V. J.; Swiderek, K. M.; Lee, T. D.; Bean,

B. P.; Adams, M. E. Nature 1992, 355, 827–829.

190. Kuhn-Nentwig, L.; Schaller, J.; Nentwig, W. Toxicon 2004, 43,







https://www.researchgate.net/publication/285840953_Excitation-Contraction_Coupling_and_Cardiac_Contractile_Force?el=1_x_8&enrichId=rgreq-5ae8bb89-6f3c-44c3-a86e-299b5b5dda46&enrichSource=Y292ZXJQYWdlOzIzMzc5OTM2NTtBUzoxMDI3NTYyODMwNjAyMjdAMTQwMTUxMDQxNTM0MA==



543–553.

191. Wullschleger, B.; Nentwig, W.; Kuhn-Nentwig, L. J Exp Biol

2005, 208, 2115–2121.

192. Branton, W. D.; Rudnick, M. S.; Zhou, Y.; Eccleston, E. D.;

Fields, G. B.; Bowers, L. D. Nature 1993, 365, 496–497.

193. Valdivia, H. H.; Kirby, M. S.; Lederer, W. J.; Coronado, R. Proc

Natl Acad Sci USA 1992, 89, 12185–12189.

194. Zamudio, F. Z.; Gurrola, G. B.; Arevalo, C.; Sreekumar, R.;

Walker, J. W.; Valdivia, H. H.; Possani, L. D. FEBS Lett 1997,

405, 385–389.

195. Lee, C. W.; Lee, E. H.; Takeuchi, K.; Takahashi, H.; Shimada,

I.; Sato, K.; Shin, S. Y.; Kim, D. H.; Kim, J. I. Biochem J 2004,

377, 385–394.

196. Fajloun, Z.; Kharrat, R.; Chen, L.; Lecomte, C.; Di Luccio, E.;

Bichet, D.; El Ayeb, M.; Rochat, H.; Allen, P. D.; Pessah, I. N.;

De Waard, M.; Sabatier, J. M. FEBS Lett 2000, 469, 179–185.

197. Esteve, E.; Mabrouk, K.; Dupuis, A.; Smida-Rezgui, S.; Altafaj,

X.; Grunwald, D.; Platel, J. C.; Andreotti, N.; Marty, I.; Sabat-

ier, J. M.; Ronjat, M.; De Waard, M. J Biol Chem 2005, 280,

12833–12839.

198. Boisseau, S.; Mabrouk, K.; Ram, N.; Garmy, N.; Collin, V.;

Tadmouri, A.; Mikati, M.; Sabatier, J. M.; Ronjat, M.; Fantini,

J.; De Waard, M. Biochim Biophys Acta 2006, 1758, 308–319.

199. Zhu, S.; Darbon, H.; Dyason, K.; Verdonck, F.; Tytgat, J.

FASEB J 2003, 17, 1765–1767.

200. Shahbazzadeh, D.; Srairi-Abid, N.; Feng, W.; Ram, N.; Borch-

ani, L.; Ronjat, M.; Akbari, A.; Pessah, I. N.; De Waard, M.; El

Ayeb, M. Biochem J 2007, 404, 89–96.

201. Schwartz, E. F.; Capes, E. M.; Diego-Garcia, E.; Zamudio, F.

Z.; Fuentes, O.; Possani, L. D.; Valdivia, H. H. Br J Pharmacol

2009, 157, 392–403.

202. Osborne, R. H. Pharmacol Ther 1996, 69, 117–142.

203. Hollmann, M.; Heinemann, S. Annu Rev Neurosci 1994, 17,

31–108.

204. Ozawa, S.; Kamiya, H.; Tsuzuki, K. Prog Neurobiol 1998, 54,

581–618.

205. Danbolt, N. C. Prog Neurobiol 2001, 65, 1–105.

206. Balcar, V. J.; Johnston, G. A. J Neurochem 1972, 19, 2657–

2666.

207. Logan, W. J.; Snyder, S. H. Brain Res 1972, 42, 413–431.

208. Amara, S. G.; Fontana, A. C. Neurochem Int 2002, 41, 313–

318.

209. Richerson, G. B.; Wu, Y. J Neurophysiol 2003, 90, 1363–1374.

210. Bruce, M.; Bukownik, R.; Eldefrawi, A. T.; Eldefrawi, M. E.;

Goodnow, R., Jr.; Kallimopoulos, T.; Konno, K.; Nakanishi, K.;

Niwa, M.; Usherwood, P. N. Toxicon 1990, 28, 1333–1346.

211. Eldefrawi, M. E.; Anis, N. A.; Eldefrawi, A. T. Arch Insect Bio-

chem Physiol 1993, 22, 25–39.

212. Mafra, R. A.; Figueiredo, S. G.; Diniz, C. R.; Cordeiro, M. N.;

Cruz, J. D.; De Lima, M. E. Brain Res 1999, 831, 297–300.

213. Oliveira, L. C.; De Lima, M. E.; Pimenta, A. M.; Mansuelle, P.;

Rochat, H.; Cordeiro, M. N.; Richardson, M.; Figueiredo, S. G.

Toxicon 2003, 42, 793–800.

214. de Figueiredo, S. G.; de Lima, M. E.; Nascimento Cordeiro,

M.; Diniz, C. R.; Patten, D.; Halliwell, R. F.; Gilroy, J.; Richard-

son, M. Toxicon 2001, 39, 309–317.

215. Massensini, A. R.; Moraes-Santos, T.; Gomez, M. V.; Romano-

Silva, M. A. Neuropharmacology 1998, 37, 289–297.

216. Lokanatha, V.; Rajendra, W. J Biochem Mol Biol Biophys

2002, 6, 167–170.

217. Libersat, F. J Comp Physiol [A] 2003, 189, 497–508.

218. Piek, T. Toxicon 1966, 4, 191–198.

219. Walther, C.; Reinecke, M. Neuroscience 1983, 9, 213–224.

220. Piek, T. J Comp Physiol [C] 1982, 72, 311–315.

221. Spanjer, W.; Grosu, L.; Piek, T. Toxicon 1977, 15, 413–421.

222. Quistad, G. B.; Nguyen, Q.; Bernasconi, P.; Leisy, D. J. Insect

Biochem Mol Biol 1994, 24, 955–961.

223. Jaques, R.; Schachter, M. Br J Pharmacol Chemother 1954, 9,

53–58.

224. Kishimura, H.; Yasuhara, T.; Yoshida, H.; Nakajima, T. Chem

Pharm Bull 1976, 24, 2896–2897.

225. Piek, T.; Schmidt, J. O.; de Jong, J. M.; Mantel, P. Comp Bio-

chem Physiol C 1989, 92, 117–124.

226. Yasuhara, T.; Mantel, P.; Nakajima, T.; Piek, T. Toxicon 1987,

25, 527–535.

227. Piek, T. Comp Biochem Physiol C 1990, 96, 223–233.

228. Piek, T.; Hue, B.; Mantel, P.; Nakajima, T.; Pelhate, M.; Yasu-

hara, T. Comp Biochem Physiol C 1990, 96, 157–162.

229. Piek, T.; Hue, B.; Mony, L.; Nakajima, T.; Pelhate, M.; Yasu-

hara, T. Comp Biochem Physiol C 1987, 87, 287–295.

230. Piek, T. Toxicon 1991, 29, 139–149.

231. Piek, T.; Hue, B.; Le Corronc, H.; Mantel, P.; Gobbo, M.; Roc-

chi, R. Comp Biochem Physiol C 1993, 105, 189–196.

232. Hue, B.; Piek, T. Comp Biochem Physiol C 1989, 93, 87–89.

233. Yoshida, H.; Geller, R. G.; Pisano, J. J. Biochemistry 1976, 15, 61–64.

234. Matsushita, N.; Miyashita, M.; Sakai, A.; Nakagawa, Y.; Miya-

gawa, H. Toxicon 2007, 50, 861–867.

235. Horita, S.; Matsushita, N.; Kawachi, T.; Ayabe, R.; Miyashita,

M.; Miyakawa, T.; Nakagawa, Y.; Nagata, K.; Miyagawa, H.;

Tanokura, M. Biochem Biophys Res Commun 2011, 411, 738–

744.

236. Silva, E. C.; Camargos, T. S.; Maranhao, A. Q.; Silva-Pereira,

I.; Silva, L. P.; Possani, L. D.; Schwartz, E. F. Toxicon 2009, 54,

252–261.

237. Shu, Q.; Liang, S. P. J Pept Res 1999, 53, 486–491.

238. Shu, Q.; Lu, S. Y.; Gu, X. C.; Liang, S. P. Acta Biochim Biophys

Sinica 2001, 33, 65–70.

239. Shu, Q.; Huang, R.; Liang, S. Eur J Biochem 2001, 268, 2301–

2307.

240. Corzo, G.; Bernard, C.; Clement, H.; Villegas, E.; Bosmans, F.;

Tytgat, J.; Possani, L. D.; Darbon, H.; Alagon, A. Biochim Bio-

phys Acta 2009, 1794, 1190–1196.

241. Vais, H.; Williamson, M. S.; Devonshire, A. L.; Usherwood, P.

N. Pest Manage Sci 2001, 57, 877–888.

242. Soderlund, D. M.; Knipple, D. C. Insect Biochem Mol Biol

2003, 33, 563–577.

243. McCutchen, B. F.; Hoover, K.; Preisler, H. K.; Betana, M. D.;

Herrmann, R.; Robertson, J. L.; Hammock, B. D. J Econ Ento-

mol 1997, 90, 1170–1180.

244. Elliott, M.; Farnham, A. W.; Janes, N. F.; Johnson, D. M.; Pul-

man, D. A.; Sawicki, R. M. Agric Biol Chem 1986, 50, 1347–

1349.

245. Georgescu, P.; Zhang, H. Nonlinear Anal: Real World Appl

2010, 11, 270–287.

246. van Frankenhuyzen, K. J Invertebr Pathol 2009, 101, 1–16.

247. Ferry, N.; Edwards, M. G.; Gatehouse, J.; Capell, T.; Christou,

P.; Gatehouse, A. M. Transgenic Res 2006, 15, 13–19.

248. Hansen, J. D.; Heidt, M. L.; Neven, L. G.; Mielke, E. A.; Bai, J.;

404 Schwartz et al.


Chen, P. M.; Spotts, R. A. Postharvest Biol Technol 2006, 40,

221–229.

249. Nagai, K.; Yano, E. Appl Entomol Zool 2000, 35, 565–574.

250. Gurr, G. M.; Kvedaras, O. L. Biol Control 2010, 52, 198–207.

251. Cory, J. S.; Hirst, M. L.; Williams, T.; Hails, R. S.; Goulson, D.;

Green, B. M.; Carty, T. M.; Possee, R. D.; Cayley, P. J.; Bishop,

D. H. L. Nature 1994, 370, 138–140.

252. Bonning, B. C.; Hammock, B. D. Annu Rev Entomol 1996, 41,

191–210.

253. Inceoglu, A. B.; Kamita, S. G.; Hinton, A. C.; Huang, Q.; Sev-

erson, T. F.; Kang, K.; Hammock, B. D. Pest Manage Sci 2001,

57, 981–987.

254. Moscardi, F. Annu Rev Entomol 1999, 44, 257–289.

255. Moscardi, F.; Souza, M. L. Biotecnol Ciencia e Desenvolvi-

mento 2002, 24, 22–29.

256. Szewczyk, B.; Hoyos-Carvajal, L.; Paluszek, M.; Skrzecz, I.;

Lobo de Souza, M. Biotechnol Adv 2006, 24, 143–160.

257. Rohrmann, G. F. Baculovirus Molecular Biology; National

Center for Biotechnology Information (US): Bethesda, MD,

2011, Chapter 9.

258. Stewart, L. M.; Hirst, M.; Lopez Ferber, M.; Merryweather, A.

T.; Cayley, P. J.; Possee, R. D. Nature 1991, 352, 85–88.

259. McCutchen, B. F.; Choudary, P. V.; Crenshaw, R.; Maddox, D.;

Kamita, S. G.; Palekar, N.; Volrath, S.; Fowler, E.; Hammock,

B. D.; Maeda, S. Biotechnology 1991, 9, 848–852.

260. Sun, X.; Chen, X.; Zhang, Z.; Wang, H.; Bianchi, F. J.; Peng,

H.; Vlak, J. M.; Hu, Z. J Invert Pathol 2002, 81, 63–69.

261. Gershburg, E.; Stockholm, D.; Froy, O.; Rashi, S.; Gurevitz,

M.; Chejanovsky, N. FEBS Lett 1998, 422, 132–136.

262. Prikhod’ko, G. G.; Popham, H. J. R.; Felcetto, T. J.; Ostlind, D.

A.; Warren, V. A.; Smith, M. M.; Garsky, V. M.; Warmke, J. W.;

Cohen, C. J.; Miller, L. K. Biol Control 1998, 12, 66–78.

263. Prikhod’ko, G. G.; Robson, M.; Warmke, J. W.; Cohen, C. J.;

Smith, M. M.; Wang, P.; Warren, V.; Kaczorowski, G.; Van der

Ploeg, L. H. T.; Miller, L. K. Biol Control 1996, 7, 236–244.

264. Hughes, P. R.; Wood, H. A.; Breen, J. P.; Simpson, S. F.; Dug-

gan, A. J.; Dybas, J. A. J Invert Pathol 1997, 69, 112–118.

265. Regev, A.; Rivkin, H.; Inceoglu, B.; Gershburg, E.; Hammock,

B. D.; Gurevitz, M.; Chejanovsky, N. FEBS Lett 2003, 537,

106–110.

266. Hughes, P. R.; Wood, H. A.; Breen, J. P.; Simpson, S. F.; Dug-

gan, A. J.; Dybas, J. A. J Invert Pathol 1997, 69, 112–118.

267. Rajendra, W.; Hackett, K. J.; Buckley, E.; Hammock, B. D. Bio-

chim Biophys Acta 2006, 1760, 158–163.

268. Pham Trung, N.; Fitches, E.; Gatehouse, J. A. BMC Biotechnol

2006, 6, 18.

269. Khan, S. A.; Zafar, Y.; Briddon, R. W.; Malik, K. A.; Mukhtar,

Z. Transgenic Res 2006, 15, 349–357.

270. Hernandez-Campuzano, B.; Suarez, R.; Lina, L.; Hernandez,

V.; Villegas, E.; Corzo, G.; Iturriaga, G. Toxicon 2009, 53, 122–

128.

271. Moser, B.; Becnel, J.; White, S.; Afonso, C.; Kutish, G.;

Shanker, S.; Almira, E. J Gen Virol 2001, 82, 283–297.

272. Gu, J. B.; Dong, Y. Q.; Peng, H. J.; Chen, X. G. Am J Trop Med

Hyg 2010, 83, 614–623.

273. Jiang, H.; Zhang, J. M.; Wang, J. P.; Yang, B.; Liu, C. F.; Lu, J.;

Hu, Y. Y. Arch Virol 2007, 152, 383–394.

274. Mukherjee, A. K.; Sollod, B. L.; Wikel, S. K.; King, G. F. Toxi-

con 2006, 47, 182–187.

275. Richardson, M.; Pimenta, A. M.; Bemquerer, M. P.; San-

toro, M. M.; Beirao, P. S.; Lima, M. E.; Figueiredo, S. G.;

Bloch, C., Jr.; Vasconcelos, E. A.; Campos, F. A.; Gomes, P.

C.; Cordeiro, M. N. Comp Biochem Physiol C 2006, 142,

173–187.

276. Kopeyan, C.; Mansuelle, P.; Martin-Eauclaire, M. F.; Rochat,

H.; Miranda, F. Nat Toxins 1993, 1, 308–312.

277. De Lima, M. E.; Martin, M. F.; Diniz, C. R.; Rochat, H. Bio-

chem Biophys Res Commun 1986, 139, 296–302.

278. Zlotkin, E.; Fishman, Y.; Elazar, M. Biochimie 2000, 82, 869–

881.

279. Escoubas, P.; Stankiewicz, M.; Takaoka, T.; Pelhate, M.; Romi-

Lebrun, R.; Wu, F. Q.; Nakajima, T. FEBS Lett 2000, 483, 175–

180.

280. Borchani, L.; Mansuelle, P.; Stankiewicz, M.; Grolleau, F.; Ces-

tele, S.; Karoui, H.; Lapied, B.; Rochat, H.; Pelhate, M.; el

Ayeb, M. Eur J Biochem 1996, 241, 525–532.

281. Cestele, S.; Borchani, L.; El Ayeb, M.; Rochat, H. FEBS Lett

1997, 405, 77–80.

282. Mejri, T.; Borchani, L.; Srairi-Abid, N.; Benkhalifa, R.; Cestele,

S.; Regaya, I.; Karoui, H.; Pelhate, M.; Rochat, H.; El Ayeb, M.

Toxicon 2003, 41, 163–171.

283. Cestele, S.; Kopeyan, C.; Oughideni, R.; Mansuelle, P.; Granier,

C.; Rochat, H. Eur J Biochem 1997, 243, 93–99.

284. Pimenta, A. M.; Martin-Eauclaire, M.; Rochat, H.; Figueiredo,

S. G.; Kalapothakis, E.; Afonso, L. C.; De Lima, M. E. Toxicon

2001, 39, 1009–1019.

285. Ali, S. A.; Stoeva, S.; Grossmann, J. G.; Abbasi, A.; Voelter, W.

Arch Biochem Biophys 2001, 391, 197–206.

286. Zlotkin, E.; Kadouri, D.; Gordon, D.; Pelhate, M.; Martin, M.

F.; Rochat, H. Arch Biochem Biophys 1985, 240, 877–887.

287. Kozminsky-Atias, A.; Somech, E.; Zilberberg, N. FEBS Lett

2007, 581, 2478–2484.







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Arthropod venoms: a vast arsenal of insecticidal neuropeptides

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