Mapping possible interaction sites for crotoxin in CNF, a gamma PLA2 inhibitor from Crotalus durissus terrificus rattle snake, using SPOT synthesis

Patricia Cota Campos; Hamine Cristina de Oliveira; Paula Ladeira Ortolani; Lutiana Amaral de Melo; Marcos R M Fontes; Consuelo Latorre Fortes-Dias

doi:10.1016/j.toxicon.2023.107267

Mapping possible interaction sites for crotoxin in CNF, a gamma PLA₂ inhibitor from Crotalus durissus terrificus rattle snake, using SPOT synthesis

Toxicon. 2023 Oct:234:107267. doi: 10.1016/j.toxicon.2023.107267. Epub 2023 Sep 1.

Authors

Patricia Cota Campos¹, Hamine Cristina de Oliveira², Paula Ladeira Ortolani¹, Lutiana Amaral de Melo¹, Marcos R M Fontes³, Consuelo Latorre Fortes-Dias⁴

Affiliations

¹ Diretoria de Pesquisa e Desenvolvimento, Fundação Ezequiel Dias, Belo Horizonte, Minas Gerais, Brazil.
² Departamento de Biofísica e Farmacologia, Instituto de Biociências, Universidade Estadual Paulista (UNESP), Botucatu, São Paulo, Brazil.
³ Departamento de Biofísica e Farmacologia, Instituto de Biociências, Universidade Estadual Paulista (UNESP), Botucatu, São Paulo, Brazil; Instituto de Estudos Avançados do Mar (IEAMar), Universidade Estadual Paulista (UNESP), São Vicente, São Paulo, Brazil.
⁴ Diretoria de Pesquisa e Desenvolvimento, Fundação Ezequiel Dias, Belo Horizonte, Minas Gerais, Brazil. Electronic address: consuelo.latorre@funed.mg.gov.br.

PMID: 37661064
DOI: 10.1016/j.toxicon.2023.107267

Abstract

Phospholipases A₂ (PLA₂s) are main components of snake venoms. Several snake species possess endogenous PLA₂ inhibitors in their circulating blood, which are generally known as sbPLIs (an acronym for snake blood phospholipase A₂inhibitors). The sbPLIs are categorized in three classes (alpha, beta or gamma) depending on the existence of distinguishing protein domains in their structure. The Crotalus durrissus terrificus venom has a highly neurotoxic PLA₂ - crotoxin (CTX) - in its composition and the self-protection of the snake is mainly ensured by a sbγPLI named CNF (standing for Crotalusneutralizing factor). In an attempt to find smaller molecules able to inhibit the catalytic activity of CTX, in the present study we used linear peptide arrays to identify CNF segments possibly involved in the interaction with the toxin. Five reacting segments were identified as possible interacting regions. The target peptides were synthesized and located in the in silico CNF structure. Although all of them are exposed to the solvent, high concentrations were needed to inhibit the PLA₂ activity of the whole venom or CTX. Limitations of the methodology employed and particular characteristics of CTX inhibition by CNF are discussed.

Keywords: CNF; Crotalus; Crotoxin; Phospholipase A(2); Phospholipase A(2) inhibitor; SPOT synthesis.