A new variant of the Ntn hydrolase fold revealed by the crystal structure of L-aminopeptidase D-Ala-esterase/amidase from Ochrobactrum anthropi

Background: The L-aminopeptidase D-Ala-esterase/amidase from Ochrobactrum a nthropi (DmpA) releases the N-terminal L and/or D-Ala residues from peptide substrates. This is the only known enzyme to liberate N-terminal amino aci ds with both D and L stereospecificity. The DmpA active form is an alpha be ta heterodimer, which results from a putative autocatalytic cleavage of an inactive precursor polypeptide. Results: The crystal structure of the enzyme has been determined to 1.82 An gstrom resolution using the multiple isomorphous replacement method. The he terodimer folds into a single domain organised as an alpha beta beta alpha sandwich in which two mixed beta sheets are flanked on both sides by two a helices; Conclusions: DmpA shows no similarity to other known aminopeptidases in eit her fold or catalytic mechanism, and thus represents the first example of a novel family of aminopeptidases. The protein fold of DmpA does, however, s how structural homology to members of the N-terminal nucleophile (Ntn) hydr olase superfamily. DmpA presents functionally equivalent residues in:the ca talytic centre when compared with other Ntn hydrolases, and is therefore li kely to use the same catalytic mechanism. In spite of this homology, the di rection and connectivity of the secondary structure elements differ signifi cantly from the consensus Ntn hydrolase topology. The DmpA structure thus c haracterises a new subfamily, but supports the common catalytic mechanism f or these enzymes suggesting an evolutionary relationship.