Purification and characterization of dog liver glucokinase

Glucokinase from dog liver has been purified to homogeneity by a procedure involving DEAE-cellulose chromatography, ammonium sulfate fractionation, gel filtration chromatography, and affinity chromatography on glucosamine-Sepharose. The purified enzyme was characterized with respect to stability, molecular weight, amino acid composition, SH groups, and physicochemical and kinetic properties. A molecular weight of 49,000 and 47,000 was estimated by sodium dodecylsulfate gel electrophoresis and gel filtration in non-denaturing conditions, respectively, indicating a monomeric structure for the enzyme. Glucokinase exhibits a sigmoidal saturation function for glucose with a Hill coefficient of 1.5 and a half-saturation value of 4mM at pH 7.5.