Biochemical characterization and pharmacological properties of a phospholipase A2 myotoxin inhibitor from the plasma of the snake Bothrops asper

Abstract

A protein that neutralizes the biological activities of basic phospholipase A2 (PLA2) myotoxin isoforms from the venom of the snake Bothrops asper was isolated from its blood by affinity chromatography with Sepharose-immobilized myotoxins. Biochemical characterization of this B. asper myotoxin inhibitor protein (BaMIP) indicated a subunit molecular mass of 23-25 kDa, an isoelectric point of 4, and glycosylation. Gel-filtration studies revealed a molecular mass of 120 kDa, suggesting that BaMIP possesses an oligomeric structure composed of five 23-25 kDa subunits. Functional studies indicated that BaMIP inhibits the PLA2 activity of B. asper basic myotoxins I and III, as well as the myotoxicity and edema-forming activity in vivo and cytolytic activity in vitro towards cultured endothelial cells, of all four myotoxin isoforms (I-IV) tested. Sequence analysis of the first 63 amino acid residues from the N-terminus of BaMIP indicated more than 65% sequence similarity to the PLA2 inhibitors isolated from the blood of the crotalid snakes Trimeresurus flavoviridis and Agkistrodon blomhoffii siniticus. These inhibitors also share sequences similar to the carbohydrate-recognition domains of human and rabbit cellular PLA2 receptors, suggesting a common domain evolution among snake plasma PLA2 inhibitors and mammalian PLA2 receptors. Despite this similarity, this is the first description of a natural anti-myotoxic factor from snake blood.

Purification and inhibitory profile of phospholipase A2 inhibitors from Australian elapid sera

“…These PLIs have been purified from Naja naja kaouthia (Thailand cobra [23,24]), Laticauda semifasciata (Sea Krait [25]), Crotalus durissus terrificus (South American rattlesnake [26][27][28]), Trimeresurus fla o iridis (Habu [29][30][31][32]), Agkistrodon blomhoffii siniticus (Chinese mamushi [33,34]) and Bothrops asper (fer-delance [35]). Additionally, an inhibitory serum factor from Vipera palaestinae (Palaestinian viper) that shares many of the characteristics of PLIs has been identified [36] ; however, it has not been clearly shown that this protein inhibits PLA # enzymes.…”

Caracterização funcional e estrutural de inibidores de fosfolipases A2 isolados do plasma de serpente Bothrops jararacussu

“…Os PLIs do tipo α têm sido encontrados na família Viperidae e Colubridae em diferentes espécies de serpentes tais como Trimeresurus flavoviridis (KOGAKI et al, 1989), Agkistrodon blomhoffii siniticus (OHKURA et al, 1993), Bothrops asper (LIZANO et al, 1997), Cerrophidion godmani , Bothrops moojeni , Gloydius brevicaudus (OKUMURA et al, 2005),…”

“…Os αPLIs são constituídos por mais de duas subunidades e apresentam uma região de domínio de reconhecimento de carboidrato (DRC) (DRICKAMER, 1988), que está presente em outras proteínas tais como a apoproteína surfactante pulmonar (WHITE et al, 1985), a proteína A ligante de manose de fígado de rato (DRICKAMER; DORDAL; REYNOLDS, 1986) e receptores de PLA 2 s de mamíferos (CUPILLARD et al, 1999 LIZANO et al, 1997;.…”

“…Considerando-se que vários PLIs estão presentes no sangue de diferentes espécies de serpentes (NOBUHISA et al, 1997;OHKURA et al, 1997;PERALES, 2003) e inibem PLA 2 s tóxicas específicas dentro da mesma espécie (LIZANO et al, 1997;, é possível sugerir que proteínas endógenas de serpentes podem estar envolvidas na neutralização de atividades enzimáticas, farmacológicas e tóxicas. -dependentes, como por exemplo, a apoproteína surfactante do pulmão, que liga os resíduos do carboidrato que contêm manose (INOUE et al, 1991;OHKURA et al, 1993;LIZANO et al, 1997).…”

“…-dependentes, como por exemplo, a apoproteína surfactante do pulmão, que liga os resíduos do carboidrato que contêm manose (INOUE et al, 1991;OHKURA et al, 1993;LIZANO et al, 1997).…”