The 2,3-bisphosphoglycerate-independent phosphoglycerate mutase from Trypanosoma brucei: metal-ion dependency and phosphoenzyme formation.

Recombinant cofactor-independent phosphoglycerate mutase from Trypanosoma brucei was inactivated by EDTA, and reactivated by Co(2+) much more than by Mn(2+) or Fe(2+). It displayed a minor phosphoglycerate phosphatase activity, which was stimulated by Mn(2+) more than by Co(2+). Upon incubation with [(32)P]phosphoglycerate, radioactivity was incorporated into the enzyme, most particularly in the presence of Mn(2+) or Fe(2+). The phosphorylated residue was identified by tandem mass spectrometry as Ser74, a residue homologous to the phosphorylated serine in alkaline phosphatase. However, the rates of formation and of disappearance of this phosphoenzyme were quite low compared to the mutase reaction. This and other properties indicated that the observed phosphoenzyme is an intermediate in the minor phosphatase activity rather than in the phosphomutase reaction.