Isolation of a phosphoryl choline-binding protein from the hemolymph of the snail, Achatina fulica

A phosphorylcholine-binding protein from the hemolymph of the snail Achatina fulica was purified to near homogeneity using a Sepharose phenylphosphorylcholine affinity column. The protein bound to the affinity column was eluted with 5 mM phosphorylcholine as a single symmetrical peak. The purified protein (400 Kda) contained 35–40% carbohydrate. On SDS-PAGE the protein separated into two bands of 20 and 24 Kda, and had a pI of 5.9. On immunodiffusion, antiserum to the snail phosphorylcholine binding protein did not cross-react against other phosphorylcholine binding proteins, like rat serum phosphorylcholine-binding protein (PCBP), limulus C-reactive protein (CRP), or human CRP. On pretreatment of the snail hemolymph with this antiserum, the hemagglutination titer of the hemolymph was markedly decreased. The purified snail phosphorylcholine binding protein agglutinated rabbit erythrocytes in the absence of divalent cation (Ca+2) but trace amount of Ca+2 increased its binding. The strongest inhibitor of the agglutination reaction was lactose, followed by melibiose and 2-deoxygalactose. The relationships of the snail phosphorylcholine binding protein to other hemolymph agglutinins and to CRPs are discussed in light of common phylogeny.

Characterization of Major Zinc Containing Myonecrotic and Procoagulant Metalloprotease `Malabarin' from Non Lethal Trimeresurus malabaricus Snake Venom with Thrombin Like Activity: Its Neutralization by Chelating Agents

A major myonecrotic zinc containing metalloprotease `malabarin' with thrombin like activity was purified by the combination of gel permeation and anion exchange chromatography from T. malabaricus snake venom. MALDI-TOF analysis of malabarin indicated a molecular mass of 45.76 kDa and its N-terminal sequence was found to be Ile-Ile-Leu-Pro(Leu)-Ile-Gly-Val-Ile-Leu(Glu)-Thr-Thr. Atomic absorption spectral analysis of malabarin raveled the association of zinc metal ion. Malabarin is not lethal when injected i.p. or i.m. but causes extensive hemorrhage and degradation of muscle tissue within 24 hours. Sections of muscle tissue under light microscope revealed hemorrhage and congestion of blood vessel during initial stage followed by extensive muscle fiber necrosis with elevated levels of serum creatine kinase and lactate dehydrogenase activity. Malabarin also exhibited strong procoagulant action and its procoagulant action is due to thrombin like activity; it hydrolyzes fibrinogen to form fibrin clot. The enzyme preferentially hydrolyzes A alpha followed by B beta subunits of fibrinogen from the N-terminal region and the released products were identified as fibrinopeptide A and fibrinopeptide B by MALDI. The myonecrotic, fibrinogenolytic and subsequent procoagulant activities of malabarin was neutralized by specific metalloprotease inhibitors such as EDTA, EGTA and 1, 10-phenanthroline but not by PMSF a specific serine protease inhibitor. Since there is no antivenom available to neutralize local toxicity caused by T. malabaricus snakebite, EDTA chelation therapy may have more clinical relevance over conventional treatment.

The sequence and structure of snake gourd (Trichosanthes anguina) seed lectin, a three-chain nontoxic homologue of type II RIPs

The sequence and structure of snake gourd seed lectin (SGSL), a nontoxic homologue of type II ribosome-inactivating proteins (RIPs), have been determined by mass spectrometry and X-ray crystallography, respectively. As in type II RIPs, the molecule consists of a lectin chain made up of two beta-trefoil domains. The catalytic chain, which is connected through a disulfide bridge to the lectin chain in type II RIPs, is cleaved into two in SGSL. However, the integrity of the three-dimensional structure of the catalytic component of the molecule is preserved. This is the first time that a three-chain RIP or RIP homologue has been observed. A thorough examination of the sequence and structure of the protein and of its interactions with the bound methyl-alpha-galactose indicate that the nontoxicity of SGSL results from a combination of changes in the catalytic and the carbohydrate-binding sites. Detailed analyses of the sequences of type II RIPs of known structure and their homologues with unknown structure provide valuable insights into the evolution of this class of proteins. They also indicate some variability in carbohydrate-binding sites, which appears to contribute to the different levels of toxicity exhibited by lectins from various sources.

Seasonal variability of rainfall recorded in growth bands of the Giant African Land Snail Lissachatina fulica (Bowdich) from India

Seasonal rainfall patterns in Bangalore, India, have been reconstructed using stable isotopic ratios in the growth bands of Giant African Land Snail shells. The present study was conducted at Bangalore, India which receives rain during the summer months. The oxygen isotopic record in the rainwater samples collected during different months covering the period of the summer monsoon of the year 2008 is compared with the isotopic ratio in the gastropod growth bands deposited simultaneously. The chronology of the shell growth band is independently established assuming the growth rate observed in a chamber experiment maintaining similar relative humidity and temperature conditions. A consistent pattern observed in the isotopic ratio in the gastropod growth bands and rainwater is demonstrated and provides a novel approach for precipitation reconstruction at seasonal and weekly time scales. This approach of using isotopic ratios in the gastropod growth bands for rainfall can serve as a substitute for filling gaps in rainfall data and for cases where no rain records are available. In addition, they can be used to determine the frequencies and magnitudes of dry spells from the past records. (C) 2013 Elsevier B.V. All rights reserved.

FURTHER RECORDS OF THE SINDH AWL-HEADED SNAKE, Lytorhynchus paradoxus (GUNTHER, 1875), FROM INDIA WITH NOTES ON ITS HABITAT AND NATURAL HISTORY

This paper presents additional distributional records of the Sindh awl-headed snake Lytorhynchus paradoxus from India, along with scale counts, measurements and natural history observations of this poorly known species.

Novel M-Superfamily and T-Superfamily conotoxins and contryphans from the vermivorous snail Conus figulinus

The venom of Conus figulinus, a vermivorous cone snail, found in the south east coast of India, has been studied in an effort to identify novel peptide toxins. The amino acid sequences of seven peptides have been established using de novo mass spectrometric based sequencing methods. Among these, three peptides belong to the M-Superfamily conotoxins, namely, Fi3a, Fi3b, and Fi3c, and one that belongs to the T-Superfamily, namely, Fi5a. The other three peptides are contryphans, namely, contryphans fib, fic, and fid. Of these Fi3b, Fi3c, Fi5a, and contryphan fib are novel and are reported for the first time from venom of C.figulinus. The details of the sequencing methods and the relationship of these peptides with other M'-Superfamily conotoxins from the fish hunting and mollusk hunting clades are discussed. These novel peptides could serve as a lead compounds for the development of neuropharmacologically important drugs. Copyright (c) 2014 European Peptide Society and John Wiley & Sons, Ltd.

A sleep-inducing peptide from the venom of the Indian cone snail Conus araneosus

The marine snail Conus araneosus has unusual significance due to its confined distribution to coastal regions of southeast India and Sri Lanka. Due to its relative scarceness, this species has been poorly studied. In this work, we characterized the venom of C. araneosus to identify new venom peptides. We identified 14 novel compounds. We determined amino acid sequences from chemically-modified and unmodified crude venom using liquid chromatography-electrospray ionization mass spectrometry and matrix assisted laser desorption ionization time-of-flight mass spectrometry. Ten sequences showed six Cys residues arranged in a pattern that is most commonly associated with the M-superfamily of conotoxins. Four other sequences had four Cys residues in a pattern that is most commonly associated with the T-superfamily of conotoxins. The post-translationally modified residue (pyroglutamate) was determined at the N-terminus of two sequences, ar3h and ar3i respectively. In addition, two sequences, ar3g and ar3h were C-terminally amidated. At a dose of 2 nmol, peptide ar3j elicited sleep when injected intraperitoneally into mice. To our knowledge, this is the first report of a peptide from a molluscivorous cone snail with sleep-inducing effects in mice. The novel peptides characterized herein extend the repertoire of unique peptides derived from cone snails and may add value to the therapeutic promise of conotoxins. (C) 2015 Elsevier Ltd. All rights reserved.