The DmpA aminopeptidase from Ochrobactrum anthropi LMG7991 is the prototype of a new terminal nucleophile hydrolase family

The DmpA (D-aminopeptidase A) protein produced by Ochrobactrum anthropi hyd rolyses p-nitroanilide derivatives of glycine and D-alanine more efficientl y than that of L-alanine. When regular peptides are utilized as substrates, the enzyme behaves as an aminopeptidase with a preference for N-terminal r esidues in an L configuration, thus exemplifying an interesting case of ste reospecificity reversal. The best-hydrolysed substrate is L-Ala-Gly-Gly, bu t tetra- and penta-peptides are also efficiently hydrolysed. The gene encod es a 375-residue precursor, but the active enzyme contains two polypeptides corresponding to residues 2-249 (alpha-subunit) and 250-375 (beta-subunit) of the precursor. Residues 249 and 250 are a Gly and a Ser respectively, a nd various substitutions performed by site-directed mutagenesis result in t he production of an uncleaved and inactive protein. The N-terminal Ser resi due of the beta-subunit is followed by a hydrophobic peptide, which is pred icted to form a beta-strand structure. All these properties strongly sugges t that DmpA is an N-terminal amidohydrolase. An exploration of the database s highlights the presence of a number of open reading frames encoding relat ed proteins in various bacterial genomes. Thus DmpA is very probably the pr ototype of an original family of N-terminal hydrolases.