Characterization of a novel phenylalanine-specific aminopeptidase from Schizophyllum commune

An aminopeptidase (APF1) with high specificity for phenylalanine and tyrosine in the active site P-1 position was purified from mycelial extracts of the wood-decaying basidiomycete Schizophyllum commune. A two-dimensional preparative electrophoretic scheme resulted in a 60-fold purification of the enzyme, which was found to have an M(r) of approximately 60 kDa by size exclusion chromatography. SDS-PAGE resolved two closely migrating subunits of approximately 30 and 31 kDa. Its pi was found to be 5.5 and its activity optimum was pH 6.5. Of 11 aminoacyl-p-nitroanilide (-pNA) substrates tested, activity was found only against Phe-pNA. APF1 also showed activity against Phe-beta-naphthylamide (-beta NA) and Tyr-beta NA; however, the activity against Tyr-beta NA was less than one third of that against Phe-beta NA. No other AA-beta NA substrates were hydrolysed. APF1. hydrolysed a variety of dipeptides with Phe in the P-1 position; the amino acid in the P-1' position affected the rate. No activity was found against dipeptides where Phe was not the N-terminus nor against substrates that were N-blocked. APF1 was partially inhibited by 1,10-phenanthroline, and this inhibition was reversible by the addition of Zn2+.