||A Porphyromonas gingivalis Periplasmic Novel Exopeptidase, Acylpeptidyl Oligopeptidase, Releases N-Acylated Di- and Tripeptides from Oligopeptides
Nemoto, Takayuki K ,
Ohara-Nemoto, Yuko ,
Bezerra, Gustavo Arruda ,
Shimoyama, YuKimura, Shigenobu
Journal of Biological Chemistry
5925 , 2016-03-11 , American Society for Biochemistry and Molecular Biology Inc.
Exopeptidases including dipeptidyl- and tripeptidyl-peptidase are crucial for the growth of Porphyromonas gingivalis, a periodontopathic asaccharolytic bacterium that incorporates amino acids mainly as di- and tri-peptides. In this study, we identified a novel exopeptidase, designated acylpeptidyl oligopeptidase (AOP), composed of 759 amino acid residues with active Ser615 and encoded by PGN_1349 in P. gingivalis ATCC 33277. AOP is currently listed as an unassigned S9-family peptidase or prolyl oligopeptidase. Recombinant AOP did not hydrolyze a Pro-Xaa bond. In addition, though sequence similarities to human and archaea-type acylaminoacyl peptidase sequences were observed, its enzymatic properties were apparently distinct from those, as AOP scarcely released an N-acyl-amino acid as compared to di- and tri-peptides, especially with N-terminal modification. The kcat/Km value against benzyloxycarbonyl-Val-Lys- Met-4-methycoumaryl-7-amide, the most potential substrate, was 123.3 ± 17.3 µM-1sec-1, optimal pH was 7-8.5, and the activity was decreased with increased NaCl concentrations. AOP existed predominantly in the periplasmic fraction as a monomer, while equilibrium between monomers and oligomers was observed with a recombinant molecule, suggesting a tendency of oligomerization mediated by the N-terminal region (Met16-Glu101). The three dimensional modeling revealed the three domain structures: residues Met16-Ala126, which has no similar homologue with known structure, residues Leu127-Met495 (β-propeller domain) and residues Ala496-Phe736 (α/β hydrolase domain), and further indicated the hydrophobic S1 site of AOP in accord with its hydrophobic P1 preference. AOP orthologues are widely distributed in bacteria, archaea, and eukaryotes, suggesting its importance for processing of nutritional and/or bioactive oligopeptides.