Contribution of W229 to the transglycosylation activity of 4-alpha-glucanotransferase from Pyrococcus furiosus |
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Authors: | Tang Shuang-Yan Yang Sung-Jae Cha Hyunju Woo Eui-Jeon Park Cheonseok Park Kwan-Hwa |
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Institution: | Center for Agricultural Biomaterials and Department of Food Science and Biotechnology, School of Agricultural Biotechnology, Seoul National University, Seoul 151-742, Korea. |
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Abstract: | A W229H mutant of 4-alpha-glucanotransferase (4-alpha-GTase) from Pyrococcus furiosus was constructed and its catalytic properties were studied to investigate the role of W229 in the catalytic specificities of the enzyme. Various activities and kinetic parameters were determined for the wild-type and W229H mutant enzymes. The transglycosylation factor and transglycosylation activity of the mutant enzyme markedly decreased, but its hydrolysis activity was scarcely affected. It was discovered that the k(cat)/K(m) value of transglycosylation activity significantly decreased to about 15% of that of the wild type, while k(cat)/K(m) value of hydrolysis activity changed little for the mutant enzyme. The hydrophobicity of W229 was thought to be critical to the transglycosylation activity of the enzyme based on the enzyme's modeled tertiary structures. |
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