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Characteristics of protein insectotoxin from the venom of the black widow spider
Ninth European Symposium
29 3
group of proteins. Then the classifying scores were calculated for each group. An example of such analysis is given in the table below where the ranges of relative content of the four amino acids in the protein tensed G, C, W, Q are given together with the corresponding classifying scores for three types toxins .
Features
Intervals
Glycine G
G ~ 5, 4 5,48,8 C 5 12, 7 12,7 13, 3 W,1 1l,7
Cysteine C Tryptophan W Glutamine O
Long neurotoxin
Short neurotoxin
Cytoxin
l2 23 0 6 l 29 2 22 0 25 26 15
0 23 12
26 1 0
O 5 3, 1 O>3, l
3 22 20 0 3 24 8 0 30
0 25 0 23 0 16 0 27 0
The appropriate scores are summarized and their maximal sum shows to which class the toxin belongs. Using this method, we have analysed a number of toxins outside of the training sampling extracted from sea snakes' venoms and anemone. We have determined that they are neurotoxins . For example we selected the toxin RTX-I from the anemone Radianthrrs macrodactylus. Its most informative features are given in the table below with their scores .
Features
LNT
SNT
CT
G = 6, 2 C=12,4 W=1,7 Q = I, 7
23 6 0 26
23 3 24 0
1 0 16 27
Thus that is a long neurotoxin . This method can be used for other compounds. Characteristics of protein insectotoxin from the venom of the black widow spider . V. N.
PASHROV, G. I. SIId O. V. Bur.c~rcov (Branch of M . M. Shemyakin Institute of Bioorganic Chemistry, U.S .S.R . Academy of Sciences, Pushchino, Moscow Region, U.S.S .R.) . KOVALEVSKAYA
Try rxoT~eix insectotoxin of presynaptic action (further one-latroinsectotoxin-LITx) was isolated from the venom of spider Latrodectus mactans tredecimgeettatus, exhausted on a column with monoclonal antibodies against a-latrotoxin (LTx). Ion-exchange and hydrophobic chromatography techniques were used (Kov~vsruYe et al., 1990). LITx was obtained in the form of a homogeneous preparation containing high-molecular polypeptide of approximately I20kDa mol. wt according to data of SDS-gradient gel electrophoresis. From plots of the equilibrium binding data, it could be ascertained, that nerve membranes prepared from the head ganglia of Gryllades suplicans contain saturable binding sites for '~I-LITx with Ilb 3 x 10 - '° M and B~ 0.07 pmole/mg membrane protein. LITx and LTx have the same N-terminal amino acid residue (glutamic acid) but a different N-terminal sequence of amino acid and differs in amino acid composition . At isoelectrofocusing both the toxins are represented by several closely-situated polypeptides with iscelectric point about 5.3 . Monospecific polyclonal antibodies against LTx in immunoeloctroblotting assays did not react with LITx . REFERENCE KOVALEVSIUYA,
G. I. et al. Bioorgan. Chem . (1990) 16, (in press).
29 3
group of proteins. Then the classifying scores were calculated for each group. An example of such analysis is given in the table below where the ranges of relative content of the four amino acids in the protein tensed G, C, W, Q are given together with the corresponding classifying scores for three types toxins .
Features
Intervals
Glycine G
G ~ 5, 4 5,4
Cysteine C Tryptophan W Glutamine O
Long neurotoxin
Short neurotoxin
Cytoxin
l2 23 0 6 l 29 2 22 0 25 26 15
0 23 12
26 1 0
O 5 3, 1 O>3, l
3 22 20 0 3 24 8 0 30
0 25 0 23 0 16 0 27 0
The appropriate scores are summarized and their maximal sum shows to which class the toxin belongs. Using this method, we have analysed a number of toxins outside of the training sampling extracted from sea snakes' venoms and anemone. We have determined that they are neurotoxins . For example we selected the toxin RTX-I from the anemone Radianthrrs macrodactylus. Its most informative features are given in the table below with their scores .
Features
LNT
SNT
CT
G = 6, 2 C=12,4 W=1,7 Q = I, 7
23 6 0 26
23 3 24 0
1 0 16 27
Thus that is a long neurotoxin . This method can be used for other compounds. Characteristics of protein insectotoxin from the venom of the black widow spider . V. N.
PASHROV, G. I. SIId O. V. Bur.c~rcov (Branch of M . M. Shemyakin Institute of Bioorganic Chemistry, U.S .S.R . Academy of Sciences, Pushchino, Moscow Region, U.S.S .R.) . KOVALEVSKAYA
Try rxoT~eix insectotoxin of presynaptic action (further one-latroinsectotoxin-LITx) was isolated from the venom of spider Latrodectus mactans tredecimgeettatus, exhausted on a column with monoclonal antibodies against a-latrotoxin (LTx). Ion-exchange and hydrophobic chromatography techniques were used (Kov~vsruYe et al., 1990). LITx was obtained in the form of a homogeneous preparation containing high-molecular polypeptide of approximately I20kDa mol. wt according to data of SDS-gradient gel electrophoresis. From plots of the equilibrium binding data, it could be ascertained, that nerve membranes prepared from the head ganglia of Gryllades suplicans contain saturable binding sites for '~I-LITx with Ilb 3 x 10 - '° M and B~ 0.07 pmole/mg membrane protein. LITx and LTx have the same N-terminal amino acid residue (glutamic acid) but a different N-terminal sequence of amino acid and differs in amino acid composition . At isoelectrofocusing both the toxins are represented by several closely-situated polypeptides with iscelectric point about 5.3 . Monospecific polyclonal antibodies against LTx in immunoeloctroblotting assays did not react with LITx . REFERENCE KOVALEVSIUYA,
G. I. et al. Bioorgan. Chem . (1990) 16, (in press).