Purification and properties of phosphorylase from Phymatotrichum omnivorum

Abstract The glycogen phosphorylase (EC 2.4.1.1) from the mycelium of Phymatotrichum omnivorum was purified by ammonium sulfate fractionation, gel nitration on Sephacryl S-200, and DEAE-cellulose ion-exchange chromatography to more than 100-fold. The purified enzyme was homogeneous; this was confirmed by polyacrylamide gel electrophoresis. Sodium dodecyl sulfate-gel electrophoresis indicated the relative molecular size of the enzyme was around 145,000. The approximate molecular weight by gel filtration was 116,000. The optimum pH of the enzyme was 7.0 and the enzyme was more specific for glycogen, with a K m value of 0.36 mg/ml. Nucleotides AMP, ADP, and ATP and compounds containing an “SH” group inhibited the enzyme activity. Diethyldithiocarbamate, EDTA, ethylene glycol bis(β-aminoethyl ether)- N,N′ -tetraacetic acid, and Cu 2+ were the potent inhibitors of the glycogen phosphorylase activity, Ca 2+ , Cu 2+ , Co 2+ , and Fe 2+ stimulated the enzyme activity. The enzyme preparation was stable at 4 °C during a period of 30 days.