Characterization, Gene Cloning, and Sequencing of a Fungal Phytase, PhyA, From Penicillium oxalicum PJ3

摘要

A phytase from Penicillium oxalicum PJ3, PhyA, was purified near to homogeneity with 427-fold increase in specific phytase activity by ammonium sulfate precipitation, gel filtration, and ionexchange chromatographies. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDSPAGE) and zymogram analysis of the purified enzyme indicated an estimated molecular mass of 65 kD. The optimal pH and temperature of the purified enzyme were pH 4.5 and 55°C, respectively. The enzyme activity was strongly inhibited by Ca~(2+), Cu~(2+), Zn~(2+), and phenylmethylsulfonyl fluoride (PMSF). The K_(m) value for sodium phytate was 0.545mM with a V_(max) of 600U/mg of protein. The phyA gene was cloned, and it contains an open reading frame of 1,383 with a single intron (118 bp), and encodes a protein of 461 amino acids.