Cloning and characterization of an antibacterial L-amino acid oxidase from Crotalus durissus cumanensis venom

An L-amino acid oxidase (LAAO) from Crotalus durissus cumanensis venom (CdcLAAO) was purified to homogeneity using a combination of size-exclusion and ion exchange chromatographies. CdcLAAO is a monomeric protein exhibiting an apparent molecular mass of 55 kDa and a calculated pI of 8. Its complete 498-amino-acid sequence was deduced through cDNA and protein sequencing. The enzyme oxidized L-Leu with K-m and a V-Max of 9.23 mu M and 0.46 mu M/min respectively, and exhibited Kcat and a Kcat/K-m of 1.8 s(-1) and 195 mM(-1)s(-1). CdcLAAO inhibited in a dose-dependent manner the growth of Staphylococcus aureus and Acinetobacter baumannii. The inhibitory effect was more significant on S. aureus, with a Minimal Inhibitory Concentration (MIC) of 8 mu g/mL and Minimal Bactericidal Concentration (MBC) of 16 mu g/mL, than against A. baumannii, with a MIC of 16 mu g/mL and MBC of 32 mu g/mL. However, against Escherichia coli CdcLAAO did not show inhibitory capacity at the concentrations tested (2-128 mu g/mL). CdcLAAO did not exhibit cytotoxic activity on the mouse myoblast cell line C2C12 and on peripheral blood mononuclear cell (PBMC). (C) 2012 Elsevier Ltd. All rights reserved.