Purification and Characterization of Glucokinase in Escherichia coli B |
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Authors: | Yasuki Fukuda Shotaro Yamaguchi Makoto Shimosaka Kousaku Murata Akira Kimura |
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Institution: | Research Institute for Food Science, Kyoto University, Uji, Kyoto 611, Japan |
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Abstract: | Glucokinase was purified from Escherichia coli B cells dosed with a hybrid plasmid carrying the gene for glucokinase. The enzyme was purified about 170-fold and was homogeneous on polyacrylamide gel electrophoresis. The enzyme was 49,000 in molecular weight and consisted of two subunits having a molecular weight of 24,500. The glucokinase catalyzed phosphorylation of D-glucose, D-mannose, D-glucosamine, and 2-deoxy-D-glucose, consuming ATP as a phosphoryl donor. Besides ATP, other nucleoside triphosphates such as ITP, GTP and UTP were also utilized as phosphoryl donors. The enzyme required free sulfhydryl groups and Mg2+ for activity. Other properties of the glucokinase were characterized and compared with those of glucokinases from various sources. |
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