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CHEMICAL AND FUNCTIONAL CHARACTERIZATION OF SCORPION TOXINS 
LouRivAL DoMiNGOs PossANi, Ph.D., International Research Scholar 
Scorpionism is a public health problem in Mex- 
ico. More than 200,000 people are stung by scor- 
pions annually, and about 700 die. For 15 years Dr. 
Possani and his colleagues have been studying the 
small peptides responsible for the lethality of scor- 
pion stings. 
Three distinct families of peptides have been iso- 
lated and characterized from the venom of these 
arachnids: 1) short-chain peptides of about 38-39 
amino acid residues, blockers of channels in 
various excitable tissues; 2) medium-chain peptides 
(61-65 amino acid residues), blockers of Na"^ chan- 
nels in mammalian tissues; and 3) long-chain pep- 
tides (about 70 amino acid residues), blockers of 
Na"*" channels of tissues in crustaceans and insects. 
The research conducted in the past year w^as fo- 
cused on the following aspects: further characteriza- 
tion of Na"^ channel-blocking peptides, isolated 
from the venom of the scorpion Centruroides nox- 
ius; primary structure determination of newly puri- 
fied K"^ channel toxins; and cloning of scorpion 
toxin genes. Additional work was performed on the 
pancreatic secretagogue effect of toxins purified 
from scorpion venoms, in collaboration with Dr. 
Paul Fletcher (East Carolina University, Greenville, 
North Carolina) . 
Definition of Four Distinct Epitopes 
in the Structure of Toxins from 
the Genus Centruroides 
Toxins 2 and 3, isolated from the venom of the 
Mexican scorpion Centruroides noxius Hoffmann, 
were purified and their amino acid sequences deter- 
mined (66 amino acid residues each). Sequence 
comparison indicates 79% identity in the primary 
structure of both toxins and shows a high similarity 
to previously characterized Centruroides toxins, 
the most similar toxins being toxin 1 from Centru- 
roides limpidus tecomanus and toxin 2 from Cen- 
truroides suffusus. Six monoclonal antibodies 
(mABs) were obtained and used to characterize 
these toxins immunochemically. Four different 
mABs reacted only with toxin 2, whereas the two 
other mABs reacted with both toxins 2 and 3 with 
the same affinity. 
Simultaneous binding of mAB pairs to the toxin 
and cross-reactivity of the venoms of different scor- 
pions with the mABs were examined, allowing the 
definition of four distinct epitopes (A-D) in the 
structure of toxins from the genus Centruroides. 
Epitope A is topographically unrelated to epitopes 
B, C, and D, but the latter three appear to be more 
related or in close proximity to one another. Epi- 
tope A was found in all Centruroides venoms tested 
(C. noxius, C. elegans, C. suffusus, C. I. limpidus, 
C. I. tecomanus, and C. I. acatlanensis) , as well as 
in four different purified toxins of C. noxius, and 
thus seems to correspond to a highly conserved 
structure. All six mABs inhibited the binding of 
toxin 2 to rat brain synaptosomal membranes, but 
only the BCF2 mAB, which belongs to the IgG2a sub- 
class, displayed a clear neutralizing activity in vivo. 
New Channel-blocking Toxins Purified 
from the Venom of C. I. limpidus 
Several years ago Dr. Possani 's group discovered 
noxiustoxin (NTX) , the first peptide purified from 
scorpion venom for which a clear effect on the 
blockade of channels was shown. Presently the 
research effort is being focused on the isolation and 
characterization of similar peptides from other scor- 
pion species. In this respect, a set of different clones 
producing mABs against NTX was obtained. Using an 
ELISA assay with these mABs, two new toxins were 
purified and sequenced from the venom of the Mex- 
ican scorpion C. I. limpidus. A series of distinct 
chromatographic fractions from this venom were at- 
tached to the plates and developed with the mAB 
anti-NTX. With this technique two different pep- 
tides were purified and sequenced, in collaboration 
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