Purification and properties of pi-repressible acid phosphatases from Aspergillus nidulans

Abstract

Two forms of the pacA-encoded acid phosphatase (designated acid phosphatases I and II) secreted by the mold Aspergillus nidulans grown in low-Pi medium at 37°, pH 5.0, were purified to apparent homogeneity by PAGE. The M r of the purified enzyme forms were ca 115 000 (60 000) and 113 000 (62 000) respectively for forms I and II secreted by strain biA1 and ca 118 000 (60 000) and 121 000 (61 000) respectively for forms I and II secreted by strain biA1 pacA1, as determined by exclusion chromatography (number between brackets are the M r as determined by SDS-PAGE). All of these purified enzyme forms showed an apparent optimum pH ranging from 6.0 to 6.5 and no deviation from Michaelis kinetics for the hydrolysis of both p-nitrophenylphosphate and α-naphthylphosphate. Heat inactivation at 60° and at pH 6.0 showed half-lives of 14 min ( k=0.033 min −1) and 10 min ( k=0.069 min −1), respectively, for the purified acid phosphatases I and II secreted by biA1 strain and half-lives of 0.8 min ( k=0.92 min −1) and 0.6 min ( k=0.95 min −1), respectively, for the purified forms I and II secreted by the biA1 pacA1 strain. The neutral sugar content of purified acid phosphatases I and II secreted by strain biA1 was 48% and 37% (w/w), respectively, whereas the content of forms I and II secreted by strain biA1 pacA1 was 18% and 11%, respectively.