High-yield purification of glucokinase from rat liver.
A rapid and reliable method for the purification of rat liver glucokinase was developed. The procedure consists of DEAE-cellulose ion-exchange chromatography, Phenyl-Sepharose hydrophobic interaction chromatography, DEAE-Affi Gel Blue dye-ligand chromatography, and duplicate steps of glucosamine-Sepharose affinity chromatography. Glucokinase was purified to a specific activity of 290 units/mg protein in a yield of 55% in 6 days. The final enzyme preparations were completely homogeneous in most experiments as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The estimated molecular weight (51,000) and sigmoidal saturation function for glucose of purified glucokinase were in good agreement with published data.
Miwa
I
I
Department of Clinical Biochemistry, Faculty of Pharmacy, Meijo University, Nagoya, Japan.
Mitsuyama
S
S
Toyoda
Y
Y
Murata
T
T
Okuda
J
J
D000818:Animals; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005260:Female; D005941:Glucokinase / Q000302:isolation & purification* / Q000378:metabolism; D008099:Liver / Q000201:enzymology*; D008970:Molecular Weight; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains
D005941:Glucokinase