Preparation and properties of a phosphoenzyme from the phosphoglucomutase of Micrococcus lysodeikticus

[摘要]

1. Phosphoglucomutase from Micrococcus lysodeikticus was incubated with ¹⁴C- and ³²P-labelled glucose 1,6-diphosphate and separated from the cofactor on a Sephadex column. ³²P-labelled phosphate (0.7mol/mol of enzyme) was associated with the enzyme, but no ¹⁴C label was. 2. The ³²P-labelled enzyme exchanged its label with the substrates. When the labelled enzyme was incubated in Tris buffer, pH8.3, at 30°C the proportion of exchangeable label slowly fell indicating a half-life of the phosphoenzyme of about 50h. 3. When HClO₄ was added to the labelled phosphoenzyme all of the label was precipitated with the protein and none was released as P_i. On alkaline hydrolysis P_i was released at a rate comparable with the rate of hydrolysis of the phosphoenzyme from rabbit muscle. 4. We conclude that the phosphoenzyme from Micrococcus lysodeikticus yields a relatively stable, catalytically active phosphoenzyme when treated with cofactor, and that there is no evidence for the formation of an enzyme–glucose 1,6-diphosphate complex. The properties of the phosphoenzyme, which resemble those of rabbit muscle phosphoglucomutase, suggest that the phosphate may be bound to serine.