Purification and characterization of an aminopeptidase from Lactobacillus helveticus JCM 1004

An aminopeptidase was purified to homogeneity from a cell-free extract of Lactobacillus helveticus JCM 1004 by ammonium sulfate precipitation and chromatography on DEAE-Sepharose, Sephacryl S-300 HR, HiLoad 26/60 Superdex 200pg and Mono-Q 10/10. The purified aminopeptidase had a trimeric structure and a molecular mass of similar to129 kDa. The enzyme was optimally active at pH 7.0 and 40 degreesC. The enzyme was a metallopeptidase, strongly activated by CO2+ and inhibited by Zn2+, Cu2+, Ni2+, Fe2+ and EDTA. The enzyme showed high activity toward p-nitroanilide derivatives (pNA) of amino acids and a peptide, dipeptides and tripeptides that had hydrophobic amino acids (Leu, Ala and Phe) or diaminocarboxylic acids (Lys and Arg) at the N-termini but not p-nitroanilide derivatives or peptides with proline at their N-termini or C-termini, such as Pro-pNA, Gly-Pro-pNA, Pro-Leu and Ala-Pro. (C) 2004 Elsevier Ltd. All rights reserved.