Partial purification and some properties of β-phosphoglucomutase from Lactobacillus brevis

• Published in: Archives of biochemistry and biophysics Vol. 228; no. 2; pp. 592 - 599
• Main Authors: Marechal, Luis R., Oliver, Guillermo, Veiga, Luiz A., de Ruiz Holgado, Aida A.P.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.02.1984
• Subjects: Catalysis; Chemical Phenomena; Chemistry; Enzyme Activation - drug effects; Kinetics; Lactobacillus - enzymology; Phosphoglucomutase - antagonists & inhibitors; Phosphoglucomutase - isolation & purification; Phosphoglucomutase - metabolism
• ISSN: 0003-9861; 1096-0384
• DOI: 10.1016/0003-9861(84)90027-4

A phosphoglucomutase (β-phosphoglucomutase) specific for β-glucose 1-phosphate, which catalyzes the β-glucose 1-phosphate:glucose 6-phosphate interconversion, was 560-fold purified from Lactobacillus brevis strain L 6. The isoelectric point of β-phosphoglucomutase was 3.8 and it had an apparent molecular weight of 29,000 estimated by gel chromatography. The enzyme required a divalent cation (Mn 2+ > Mg 2+ > Ni 2+ > Co 2+) and β-glucose 1,6-bisphosphate for activity. The equilibrium constant K e for the reaction β- d-glucose 1-phosphate γ d-glucose 6-phosphate at 30 °C and pH 6.7 is 18.5. β-phosphoglucomutase had a pH optimum between 6.3 and 6.8 and appeared to be quite specific: α-glucose 1-phosphate, α- or β-galactose 1-phosphate and α- or β- N-acetylglucosamine 1-phosphate did not substitute for β-glucose 1-phosphate. Double reciprocal plots of the data from initial velocity studies at five β-glucose 1-phosphate concentrations (10 to 100 μ m) and four β-glucose 1,6-bisphosphate concentrations (0.125 to 1.0 μ m) showed that the apparent Michaelis constants for β-glucose 1-phosphate and β-glucose 1,6-bisphosphate were related to the concentrations of β-glucose 1,6-bisphosphate and β-glucose 1-phosphate, respectively, in such a way as to suggest a pingpong mechanism. The same conclusion was obtained when substrate-velocity relationships were investigated at fixed ratio of both substrates: the Lineweaver-Burk plots showed linear lines and no parabolic ones. The “true” K m for β-glucose 1-phosphate and β-glucose 1,6-bisphosphate were found to be about 12 and 0.8 μ m, respectively.