Analysis of the protein profile of the venoms of snakes Bothrops asper, Bothrocophias myersi and Crotalus durissus from the Colombian Andean Region obtained by RP-HPLC

Stefano Scovino Loboguerrero,Carlos Galvis,Ana Lucía Castiblanco,Karen Sarmiento,Fabio Aristizabal

doi:10.15446/rev.colomb.biote.v23n1.94211

Abstract

Snake venoms comprise a highly complex mixture of proteins, and there is also a high interspecific and intraspecific variability in their composition, even in the same region. Our aim was to compare the composition of the venoms of Bothrocophias myersi, Crotalus durissus and Bothrops asper, snakes from the Andean region in Colombia by Reverse-Phase High-Performance Liquid Chromatography (RP-HPLC). The venoms were given to the research group under an agreement with the Fundación Zoológica de Cali. The venoms pool was obtained by manual extraction, lyophilized and refrigerated. The protein found in the venoms was quantified by spectrophotometry using the Bradford and Lowry methods and direct measurement by Nanodrop®. The protein composition was stablished by RP-HPLC, using a Lichosper 100 RP, C18 column (250X4 mm) with a pore size of 5µm, as well as by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE). The highest quantity of protein was found in the venom of B. myersi (108,6 mg/mL) followed by C. durissus (78,1 mg/mL) and B. asper (74.1 mg/mL). All venoms showed bands of 15 and 50 KDa by SDS-PAGE; The most important finding is the abundance of PLA2 and svMP in the venom of B. myersi. Chromatographic analyses revealed a very similar venom composition profile, but also certain differences in toxins abundance. We conclude that the process of separating the venom proteins by RP-HPLC and SDS-PAGE are very important as a first step to know the venoms profiles, which in turn could allow medical staff to elucidate the clinical syndrome produced by snakebites.

Highlights

Snakebite is a public health problem in Colombia with approximately 5,000 cases/year at a rate of 10.2 / 10,000 and a mortality rate of around 1%
Bothrocophias myersi is a recently discovered snake from this family with six species (Salazar Valenzuela et al, 2014); it is known as cabeza de lanza, cachetona or bufadora, and its geographical distribution includes the biogeographic area of Chocó, the surroundings of the Andes Cordillera, and the upper Amazon basin
Data revealed that B. myersi obtained the highest protein concentration, whereas B

Summary

Introduction

Snakebite is a public health problem in Colombia with approximately 5,000 cases/year at a rate of 10.2 / 10,000 and a mortality rate of around 1%. The Viperidae family is the cause of 65% of snakebite poisoning (Instituto Nacional de Salud, 2018); it is considered the most aggressive and includes the genus Bothrops sp, Crotalus sp, and Lachesis sp. Bothrops sp known in Spanish-speaking countries as mapaná, equis, pudridora, cuatronarices, jergón, among others; a possible factor of confusion in snake identification of the local population, is reflected in the names; for example, Bothrops atrox and Lachesis muta which receive the same popular name surucucu in certain Amazon areas. Bothrocophias myersi is a recently discovered snake from this family with six species (Salazar Valenzuela et al, 2014); it is known as cabeza de lanza, cachetona or bufadora, and its geographical distribution includes the biogeographic area of Chocó, the surroundings of the Andes Cordillera, and the upper Amazon basin

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Results