The Deletion of Amino-Terminal Domain in Thermoactinomyces vulgaris R-47 α-Amylases : Effects of domain N on Activity, Specificity, Stability and Dimerization
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概要
- 論文の詳細を見る
Thermoactinomyces vulgaris R-47 α-amylases, TVA I and TVA II, have a domain N, which is an extra structure in the family 13 enzymes. To investigate the roles of domain N in TVAs, we constructed TVAs-△ N mutants which are deleted in domain N, and Y14,16,68A and Y41,82,95A mutants of TVA II. TVAs-△N were unstable under alkaline conditions, and their thermal stabilities were 10℃ lower than that of wild-types. The specific activities of TVAs-△N for pullulan, starch, cyclodextrins, and oligosaccharides were drastically decreased, being about 1,500- to 10,000-fold smaller than those of wild types. The K_<cat> values of Y14,16,68A and Y41,82,92A for all tested substrates were markedly decreased, and the K_m value of Y14,16,68A for α-CD and maltotriose were 25- and 3-fold larger, and that of Y41,82,92A for starch was 10-fold larger than that of the wild-type. TVA I and TVAs-△N in solution are a monomer, while TVA II is a homo-dimer, calculated by their molecular masses. These results suggest domain N in TVAs is an important structure for stabilization of enzymes recognition and hydrolysis of substrates, and dimerization of TVA II.
- 2001-02-23
著者
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神鳥 成弘
東京農工大学工学部生命工学科
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TONOZUKA Takashi
Department of Applied Biological Science, Faculty of Agriculture, Tokyo University of Agriculture an
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殿塚 隆史
東京農工大学農学部
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Yokota T
Faculty Of Agriculture Shizuoka University
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YOKOTA Takehiro
Department of Applied Biological Science, Tokyo University of Agriculture Technology
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KAMITORI Shigehiro
Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology
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SAKANO Yoshiyuki
Department of Applied Biological Science, Tokyo University of Agriculture Technology
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Yuuki T
Asahi Breweries Ltd. Ibaraki Jpn
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Tonozuka Takashi
Department Of Applied Biological Science Faculty Of Agriculture Tokyo University Of Agriculture And
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Sakano Y
Department Of Applied Biological Science Faculty Of Agriculture Tokyo University Of Agriculture And
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Sakano Yoshiyuki
Department Of Applied Biological Science Faculty Of Agriculture Tokyo University Of Agriculture And
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神鳥 成弘
Department Of Biotechnology & Life Science Tokyo University Of Agriculture & Technology
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Yokota Takao
Iss Science Project Office Institute Of Space And Astronautical Science Japan Aerospace Exploration
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Tonozuka Takashi
Forage Crop Breeding Unit National Agricultural Research Center For Kyushu Okinawa Region
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SAKANO Yoshiyuki
Department of Agricultural Chemistry, Faculty of Agriculture, Tokyo Noko University
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