Purification and characterization of Aeromonas caviae aminopeptidase possessing debittering activity

An aminopeptidase (EC 3.4.11.10) was purified to homogeneity from the culture filtrate of Aeromonas caviae T-64. The purified enzyme showed maximum activity at pH 8.5 and 50 degrees C and was inhibited by o-phenanthroline and EDTA. The M-r values estimated by MALDI-TOF mass spectrometry and SDS-PAGE were 29 500 and 31 000 Da, respectively. These results indicate that the enzyme is a monomeric metalloenzyme. The enzyme hydrolyzed the di-and tripeptides containing hydrophobic amino acid residues such as valine, isoleucine, leucine, tyrosine, and phenylalanine in the N-terminal and/or adjacent positions with high hydrolysis efficiency (K-cat/K-m). This substrate specificity was not restricted to the di-and tripeptides as the peptides longer than tripeptides were hydrolyzed at a high rate when hydrophobic amino acid residues were located in the N-terminal region.