Purification and Characterization of Phosphoglucomutase from Heterotrophic Tissues of Brassica campestris L

Two isoforms of phosphoglucomutase (PGM, EC 2.7.5.1) designated as PGM-I and PGM-II were purified from developing seeds of Brassica campestris L to their electrophoretic homogeneity. Both the forms had molecular mass of 59 kD each and were mono meric. PGM-I exhibited maximum activity at pH 7.5, while PGM-II evinced pH optima at 8.25. Both the forms exhibited hyperbolic response towards increasing concentrations of the substrate with K_m values of 0.10 mM for PGM-I and 0.12 mM for PGM-II and had absolute requirement for glucose-1,6-P₂. Fructose-1,6-P₂ and 2,3-diphosphoglyceric acid inhibited the two forms non-com-petitively, whereas, ribulose-1,5-P₂ inhibited only PGM-II, with K_i value of 0.8 mM. ATP inhibited the enzyme uncompetitively with K₁ values for 0.26 mM (PGM-I) and 0.12 mM (PGM-II). Use of group specific protease inhibitors indicated Ser, His and Cys to play significant role in catalysis. On the basis of their differential behaviour and kinetic properties, PGM-I and PGM-II may be the forms present in cytosol and leucoplasts, respectively.