Cloning and functional expression of secreted phospholipases A2 from Bothrops diporus (Yarará Chica)

YUNES QUARTINO P.J.; BARRA J.L.; FIDELIO G.D.

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

ACADEMIC PRESS INC ELSEVIER SCIENCE

Lugar: Amsterdam; Año: 2012 vol. 427 p. 321 - 325

0006-291X

AbstractBothrops diporus is a very common viper in Argentina. At present, no completesequence of secreted phospholipase A2 (sPLA2) from this snake has been reported. We havecloned two sPLA2 isoenzymes as well as a putative sPLA2-like myotoxin from venomgland. The two sPLA2 were expressed as inclusion bodies in E. coli with an N-terminal tagof ubiquitin. After in vitro renaturation and cleavage step, using an ubiquitin specificpeptidase, the recombinants exhibited sPLA2 activity when analyzed by means of Langmuirdilauroylphosphatidylcholine monolayers as substrate. Both enzymes have a similar surfacepressure-activity profile when compared with non-recombinant purified isoforms. To ourknowledge, this is the first time that analysis of optimal lateral pressure of substratemonolayers by using the surface barostat technique is performed on recombinant sPLA2s.Keywords: Bothrops diporus; snake venom; phospholipase A2; DLPC monolayer;ubiquitin