title: Cloning and characterization of an antibacterial L-amino acid oxidase from Crotalus durissus cumanensis venom.
abstract: 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 μM and 0.46 μM/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 μg/mL and Minimal Bactericidal Concentration (MBC) of 16 μg/mL, than against A. baumannii, with a MIC of 16 μg/mL and MBC of 32 μg/mL. However, against Escherichia coli CdcLAAO did not show inhibitory capacity at the concentrations tested (2-128 μg/mL). CdcLAAO did not exhibit cytotoxic activity on the mouse myoblast cell line C(2)C(12) and on peripheral blood mononuclear cell (PBMC).
authors.affiliation: Programa Ofidismo/Escorpionismo, Universidad de Antioquia, Street 62 No. 52-59, A.A. 1226 Medellín, Colombia. johana2104@gmail.com
authors.forename: Leidy Johana
authors.initials: LJ
authors.lastname: Vargas
authors.forename: Juan Carlos
authors.initials: JC
authors.lastname: Quintana
authors.forename: Jaime Andrés
authors.initials: JA
authors.lastname: Pereañez
authors.forename: Vitelbina
authors.initials: V
authors.lastname: Núñez
authors.forename: Libia
authors.initials: L
authors.lastname: Sanz
authors.forename: Juan
authors.initials: J
authors.lastname: Calvete
mesh_terms: D000150:Acinetobacter / Q000187:drug effects* / Q000254:growth & development / Q000648:ultrastructure; D000595:Amino Acid Sequence; D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D003435:Crotalid Venoms / Q000201:enzymology* / Q000235:genetics; D017839:Crotalus / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D006801:Humans; D050830:L-Amino Acid Oxidase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology*; D007963:Leukocytes, Mononuclear / Q000187:drug effects; D051379:Mice; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D032446:Myoblasts / Q000187:drug effects; D010084:Oxidation-Reduction; D017434:Protein Structure, Tertiary; D017422:Sequence Analysis, DNA; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000254:growth & development / Q000648:ultrastructure
chemical_list: D000900:Anti-Bacterial Agents; D003435:Crotalid Venoms; D018076:DNA, Complementary; D050830:L-Amino Acid Oxidase