Structural determinants of the hyperalgesic activity of myotoxic Lys49-phospholipase A2

AUTOR(ES)

Zambelli, Vanessa Olzon, Chioato, Lucimara, Gutierrez, Vanessa Pacciari, Ward, Richard John, Cury, Yara

FONTE

J. Venom. Anim. Toxins incl. Trop. Dis

DATA DE PUBLICAÇÃO

16/03/2017

RESUMO

Abstract Background Bothropstoxin-I (BthTx-I) is a Lys49-phospholipase A2 (Lys49-PLA2) from the venom of Bothrops jararacussu, which despite of the lack of catalytic activity induces myotoxicity, inflammation and pain. The C-terminal region of the Lys49-PLA2s is important for these effects; however, the amino acid residues that determine hyperalgesia and edema are unknown. The aim of this study was to characterize the structural determinants for the Lys49-PLA2-induced nociception and inflammation. Methods Scanning alanine mutagenesis in the active-site and C-terminal regions of BthTx-I has been used to study the structural determinants of toxin activities. The R118A mutant was employed as this substitution decreases PLA2 myotoxicity. In addition, K115A and K116A mutants – which contribute to decrease cytotoxicity – and the K122A mutant – which decreases both myotoxicity and cytotoxicity – were also used. The H48Q mutant – which does not interfere with membrane damage or myotoxic activity – was used to evaluate if the PLA2 catalytic site is relevant for the non-catalytic PLA2-induced pain and inflammation. Wistar male rats received intraplantar injections with mutant PLA2. Subsequently, hyperalgesia and edema were evaluated by the paw pressure test and by a plethysmometer. Native and recombinant BthTx-I were used as controls. Results Native and recombinant BthTx-I induced hyperalgesia and edema, which peaked at 2 h. The R118A mutant did not induce nociception or edema. The mutations K115A and K116A abolished hyperalgesia without interfering with edema. Finally, the K122A mutant did not induce hyperalgesia and presented a decreased inflammatory response. Conclusions The results obtained with the BthTx-I mutants suggest, for the first time, that there are distinct residues responsible for the hyperalgesia and edema induced by BthTx-I. In addition, we also showed that cytolytic activity is essential for the hyperalgesic effect but not for edematogenic activity, corroborating previous data showing that edema and hyperalgesia can occur in a non-dependent manner. Understanding the structure-activity relationship in BthTx-I has opened new possibilities to discover the target for PLA2-induced pain.

Documentos Relacionados

• Efeito nociceptivo induzido por fosfolipases A2 (FLA2 variantes Lys49 e Asp49) isoladas do veneno de serpentes Bothrops asper: caracterização dos mecanismos centrais e determinantes moleculares
• Efeito bactericida de Fosfolipases A2-Lys49: o papel da região C-terminal na atividade de Bothropstoxina-I em membranas biológicas e artificiais.
• Phospholipase A2 activity of low density lipoprotein: evidence for an intrinsic phospholipase A2 activity of apoprotein B-100.
• Subcellular characteristics of phospholipase A2 activity in the rat kidney. Enhanced cytosolic, mitochondrial, and microsomal phospholipase A2 enzymatic activity after renal ischemia and reperfusion.
• Modulation of the Activity of Secretory Phospholipase A2 by Antimicrobial Peptides