Isolation and biochemical, functional and structural characterization of a novel L-amino acid oxidase from Lachesis muta snake venom

Author(s):Show less -

Bregge-Silva, Cristiane ^[1] ; Nonato, Maria Cristina ^[1] ; de Albuquerque, Sergio ^[2] ; Ho, Paulo Lee ^[3] ; Junqueira de Azevedo, Inacio L. M. ^[3] ; Vasconcelos Diniz, Marcelo Ribeiro ^[4] ; Lomonte, Bruno ^[5] ; Rucavado, Alexandra ^[5] ; Diaz, Cecilia ^{[5, 6]} ; Maria Gutierrez, Jose ^[5] ; Arantes, Eliane Candiani ^[1]

Affiliation

^[1] Univ Sao Paulo, Dept Quim & Fis, Fac Ciencias Farmaceut Ribeirao Preto, BR-14040903 Ribeirao Preto, SP - Brazil

^[2] Univ Sao Paulo, Dept Anal Clin Toxicol & Bromatol, Fac Ciencias Farmaceut Ribeirao Preto, BR-14040903 Ribeirao Preto, SP - Brazil

^[3] Ctr Biotecnol, Inst Butantan, Sao Paulo - Brazil

^[4] Fundacao Ezequiel Dias, Ctr Pesquisa & Desenvolvimento, Belo Horizonte, MG - Brazil

^[5] Univ Costa Rica, Fac Microbiol, Inst Clodomiro Picado, San Jose - Costa Rica

^[6] Univ Costa Rica, Dept Bioquim, Escuela Med, San Jose - Costa Rica

• Document type: Journal article
• Source: Toxicon; v. 60, n. 7, p. 1263-1276, DEC 1 2012.
• Web of Science Citations: 39

Abstract

The aim of this study was the isolation of the LAAO from Lachesis muta venom (LmLAAO) and its biochemical, functional and structural characterization. Two different purification protocols were developed and both provided highly homogeneous and active LmLAAO. It is a homodimeric enzyme with molar mass around 120 kDa under non-reducing conditions, 60 kDa under reducing conditions in SDS-PAGE and 60852 Da by mass spectrometry. Forty amino acid residues were directly sequenced from LmLAAO and its complete cDNA was identified and characterized from an Expressed Sequence Tags data bank obtained from a venom gland. A model based on sequence homology was manually built in order to predict its three-dimensional structure. LmLAAO showed a catalytic preference for hydrophobic amino acids (K-m of 0.97 mmol/L with Leu). A mild myonecrosis was observed histologically in mice after injection of 100 mu g of LmLAAO and confirmed by a 15-fold increase in CK activity. LmLAAO induced cytotoxicity on AGS cell line (gastric adenocarcinoma, IC50: 22.7 mu g/mL) and on MCF-7 cell line (breast adenocarcinoma, IC50:1.41 mu g/mL). It presents antiparasitic activity on Leishmania brasiliensis (IC50: 2.22 mu g/nnL), but Trypanosoma cruzi was resistant to LmLAAO. In conclusion, LmLAAO showed low systemic toxicity but important in vitro pharmacological actions. (C) 2012 Elsevier Ltd. All rights reserved. (AU)

• FAPESP's process: 05/54855-0 - Animal toxins: structure, function and biotechnological applications
• Grantee: Suely Vilela
• Support Opportunities: Research Projects - Thematic Grants