title: Purification, sequencing and evaluation of a divergent phytase from Penicillium oxalicum KCTC6440.
abstract: A fungal strain producing high levels of phytase was purified to homogeneity from Penicillium oxalicum KCTC6440 (PhyA). The molecular mass of the purified PhyA was 65 kDa and optimal activity occurred at 55°C. The enzyme was stable in a pH range of 4.5-6.5, with an optimum performance at pH 5.5. The Km value for the substrate sodium phytate was 0.48 mM with a Vmax of 672 U/mg. The enzyme was inhibited by Ca(2+), Cu(2+), and Zn(2+), and slightly enhanced by EDTA. The PhyA efficiently released phosphate from feedstuffs such as soybean, rich bran and corn meal. The PhyA gene was cloned in two steps of degenerate PCR and inverse PCR and found to comprise 1501 bp and encode 461 amino acid residues. The enzyme was found to have only 13 amino acids differing to the known PhyA from other Penicillium sp., but has distinct enzyme characteristics. Computational analysis showed that PhyA possessed more positively charged residues in the active sites compared to other PhyA molecules, which may explain the broader pH spectrum.
authors.affiliation: Program in Bioinformatics and Integrative Biology, University of Massachusetts Medical School.
authors.forename: Bong-Hyun
authors.initials: BH
authors.lastname: Kim
authors.forename: Ji Yeon
authors.initials: JY
authors.lastname: Lee
authors.forename: Peter C W
authors.initials: PC
authors.lastname: Lee
mesh_terms: D010832:6-Phytase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008954:Models, Biological; D008970:Molecular Weight; D010407:Penicillium / Q000201:enzymology* / Q000235:genetics; D010710:Phosphates / Q000378:metabolism; D010833:Phytic Acid / Q000378:metabolism; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment; D013025:Glycine max / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D003313:Zea mays / Q000378:metabolism
chemical_list: D010710:Phosphates; D010833:Phytic Acid; D010832:6-Phytase