Trypanosoma brucei contains a 2,3-bisphosphoglycerate independent phosphoglycerate mutase

Assays of phosphoglycerate mutase (PGAM) activity in lysates of bloodstream form Trypanosoma brucei appeared not to require exogenous 2,3-bisphosphogl ycerate, thus suggesting that this protist contains an enzyme belonging to the class of cofactor-independent PGAMs. A gene encoding a polypeptide with motifs characteristic for this class of enzymes was cloned. The predicted T. brucei PGAM polypeptide contains 549 amino acids, with M-r 60 557 and pI 5.5. Comparison with 15 cofactor-independent PGAM sequences available in d atabases showed that the amino-acid sequence of the trypanosome enzyme has 59-62% identity with plant PGAMs and 29-35% with eubacterial enzymes. A low 28% identity was observed with the only available invertebrate sequence. T he trypanosome enzyme has been expressed in Escherichia coli, purified to h omogeneity and subjected to preliminary kinetic analysis. Previous studies have shown that cofactor-dependent and -independent PGAMs are not homologou s. It has been inferred that the cofactor-independent PGAMs are in fact hom ologous to a family of metalloenzymes containing alkaline phosphatases and sulphatases. Prediction of the secondary structure of T. brucei PGAM and th reading the sequence into the known crystal structure of E. coli alkaline p hosphatase (AP) confirmed this homology, despite the very low sequence iden tity. Generally, a good match between predicted (PGAM) and actual (AP) seco ndary structure elements was observed. In contrast to trypanosomes, glycoly sis in all vertebrates involves a cofactor-dependent PGAM. The presence of distinct nonhomologous PGAMs in the parasite and its human host offers grea t potential for the design of selective inhibitors which could form leads f or new trypanocidal drugs.