Kaliotoxin (KTX) is a natural peptide blocker of voltage-dependent K+ channels. The 3D structure of a truncated analogue of KTX (Fernandez et al. (1994) Biochemistry 33, 14256-14263) was determined by NMR spectroscopy and showed significant differences from structures established for other related scorpion toxins. A recent publication with the structure of the complete toxin (Aiyar et al. (1995) Neuron 15, 1169-1181) did not confirm these differences. In this communication we report NMR data for KTX at pH 3.0, 5.5 and 7.2 and the 3D structure obtained from data at pH = 5.5. Complete KTX displays a folding similar to that of other toxins with an alpha-helix and a beta-sheet linked by two disulphide bonds. The pKa of His 34 is anomalously low (4.7-5.2 depending on the buffer) owing to its interaction with two Lys residues (including the essential Lys 27), the charged N-terminus and the side chain of Met 29. Charged residues are placed symmetrically with respect to an axis that approximately coincides with one of the principal components of the moment of inertia of the toxin. His 34, which occupies a well-defined position between two conserved Cys, is located on the centre of a layer of charged groups. Positively and negatively charged residues are found at the same position in related toxins. It is suggested that electrostatic effects modulate the distances between positive charges in flexible side chains, contributing to the fine tuning of the selectivity toward different channel subclasses and that the approximate coincidence between the moment of inertia and the charge axis facilitate the approach of the toxin to the channel. The very low pKa of His 34 implies that it will be completely unprotonated at physiological pH.

Hydrogen Bonding↗

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Specificity of anti-peptide antibodies elicited against synthetic peptides mimicking conserved regions of HIV1 envelope glycoprotein.

Comparison of HIV1Bru and HIV2Rod external envelope glycoprotein sequences enabled us to select ten highly conserved peptide sequences. The corresponding peptides were chemically synthesized, then coupled to bovine serum albumin before injection in rabbits. Although all peptides were immunogenic, only antibodies directed against peptides P1 (amino acid residues 33-55), P22 (418-462), P8 (487-508) and P21 (487-534) were able to interact with significant affinity (K0.5 about 10(-6) to 10(-8) M) with the native glycoprotein by radioimmunoassay. Noteworthy was the capacity of anti-P1 antibodies to also recognize the glycoprotein of HIV2. Anti-peptide antibodies were tested for their ability to interfere with the gp120-CD4 interaction, membrane fusion and virus replication. Preincubation of gp120 with antibodies directed to the region previously described as the putative CD4-binding site, P22 (418-462), did not abolish gp120 binding to CD4-positive cells.

Amino Acid Sequence↗

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Lebetin peptides: potent platelet aggregation inhibitors.

Lebetins from Macrovipera lebetina snake venom constitute a new class of inhibitors of platelet aggregation. There are two groups of peptides: lebetin 1 (L1; 11- to 13-mer) and lebetin 2 (L2; 37- to 38-mer). The short lebetins are identical to the N-terminal segments of the longer ones. They inhibit platelet aggregation induced by various agonists (e.g. thrombin, PAF-acether or collagen). The shortest lebetin (11-mer) shows potent inhibition of rabbit (IC(50) = 7 nM) and human (IC(50) = 5 nM) platelets. They prevent collagen-induced thrombocytopenia in rats. N- and C-terminal-truncated synthetic L1gamma (sL1gamma; 11-mer) is less active in inhibiting platelet aggregation than the native peptide. Results from Ala scan studies of the sL1gamma peptide indicated that replacement of the residues (P3, G7, P8, P9 or N10) resulted in a remarkable drop in the activity, whereas replacement of residues K2, P4 or K6 by Ala resulted in enhancement of the antiplatelet activity by at least 10-fold. To examine the activity of multimeric L1gamma, several multimeric peptides were synthesized using the multiple-antigen peptide system assembled on a branched lysine core and their antiplatelet activity was evaluated in vitro. The largest multimeric peptides showed a 1,000-fold increase in antiplatelet activity.

Animals↗

PubMedDate not supplied

[Production of monoclonal antibodies. Scorpion antitoxins: characterization and molecular mechanisms of neutralization].

Two monoclonal antibodies specific for the potent toxin II of the scorpion Androctonus australis Hector have been produced. One of them shows both high affinity binding to the toxin (Kd = 0.8 nM) and in vivo and in vitro neutralizing properties. The mechanism by which the antibody neutralizes toxin binding to its receptor was shown to be of the competitive type, the epitope is overlapping or being close to the receptor binding region of the toxin. Several residues of the toxin clustered in the C-terminal region were shown likely to be part of the discontinuous epitope recognized by the antibody. The positive charge of the NH2-Lys58 seems to play a pivotal role in the binding of the toxin to both the monoclonal antibody and the sodium channel receptor.

Animals↗

PubMedDate not supplied

[Scorpion toxins: a study model of the structure-function relation of proteins].

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Action Potentials↗