The authors Juliana I. dos Santos and Mariana Cintra-Francischinelli contributed equally to this work.
Structural, functional, and bioinformatics studies reveal a new snake venom homologue phospholipase A2 class
Article first published online: 27 SEP 2010
Copyright © 2010 Wiley-Liss, Inc.
Proteins: Structure, Function, and Bioinformatics
Volume 79, Issue 1, pages 61–78, January 2011
How to Cite
dos Santos, J. I., Cintra-Francischinelli, M., Borges, R. J., Fernandes, C. A. H., Pizzo, P., Cintra, A. C. O., Braz, A. S. K., Soares, A. M. and Fontes, M. R. M. (2011), Structural, functional, and bioinformatics studies reveal a new snake venom homologue phospholipase A2 class. Proteins, 79: 61–78. doi: 10.1002/prot.22858
- Issue published online: 30 NOV 2010
- Article first published online: 27 SEP 2010
- Accepted manuscript online: 23 AUG 2010 10:33AM EST
- Manuscript Accepted: 13 AUG 2010
- Manuscript Revised: 22 JUL 2010
- Manuscript Received: 18 MAY 2010
- phospholipase A2 ;
- X-ray crystallography;
- phylogenetic analysis;
- myotube cell culture;
- calcium imaging
Phospholipases A2 (PLA2s) are enzymes responsible for membrane disruption through Ca2+-dependent hydrolysis of phospholipids. Lys49-PLA2s are well-characterized homologue PLA2s that do not show catalytic activity but can exert a pronounced local myotoxic effect. These homologue PLA2s were first believed to present residual catalytic activity but experiments with a recombinant toxin show they are incapable of catalysis. Herein, we present a new homologue Asp49-PLA2 (BthTX-II) that is also able to exert muscle damage. This toxin was isolated in 1992 and characterized as presenting very low catalytic activity. Interestingly, this myotoxic homologue Asp49-PLA2 conserves all the residues responsible for Ca2+ coordination and of the catalytic network, features thought to be fundamental for PLA2 enzymatic activity. Previous crystallographic studies of apo BthTX-II suggested this toxin could be catalytically inactive since a distortion in the calcium binding loop was observed. In this article, we show BthTX-II is not catalytic based on an in vitro cell viability assay and time-lapse experiments on C2C12 myotube cell cultures, X-ray crystallography and phylogenetic studies. Cell culture experiments show that BthTX-II is devoid of catalytic activity, as already observed for Lys49-PLA2s. Crystallographic studies of the complex BthTX-II/Ca2+ show that the distortion of the calcium binding loop is still present and impairs ion coordination even though Ca2+ are found interacting with other regions of the protein. Phylogenetic studies demonstrate that BthTX-II is more phylogenetically related to Lys49-PLA2s than to other Asp49-PLA2s, thus allowing Crotalinae subfamily PLA2s to be classified into two main branches: a catalytic and a myotoxic one. Proteins 2010. © 2010 Wiley-Liss, Inc.