1998 Fiscal Year Final Research Report Summary
The novel substrate recognition mechanism utilized by thermophilic aspartate aminotransferase
| Project/Area Number |
09680619
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional biochemistry
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| Research Institution | Osaka University |
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Principal Investigator |
KURAMITSU Seiki Osaka University, Graduate School of Science Professor, 大学院・理学研究科, 教授 (60153368)
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| Project Period (FY) |
1997 – 1998
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| Keywords | Thermus thermophilus / aminotransferase / transferase / X-ray crystallography / lysine / substrate recognition mechanism / arginine |
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| Research Abstract |
It has been believed that an enzyme is quite specific for its substrate.But aminotransferases are quite unique enzymes which can bind quite different binds of substrate.
These enzymes have two different types of binding pockets for acidic and hydrophobic substrates. (Catalytic residue is identical for each substrates.) Acidic substrate binds rigid region (Kawaguchi et al.(1997) J.Biochem.122, 55-63) and hydrophobic substrate binds flexible region (Kawaguchi and Kuramitsu, (1998) J.Biol. Chem. 273, 18353-18364).
The three-dimensional structure of aspartate aminotransferase from extreme thermophile, Thermus thermophilus HB8 have been determined by X-ray crystallography.In view of the X-ray crystallographic structure of T.thermophilus AspAT, K1O9V mutant was prepared.Replacing K109 with Val resulted in loss of activity toward acidic substrates, but increased that toward the neutral substrate, alanine, considerably.These results indicate that K109 is a major determinant of the acidic substrate specificity of T.thermophilus AspATs. Kinetic analysis of the interactions with neutral substrates indicated that T.thermophilus AspAT is subject to less steric hindrance and its substrate-binding pocket has a more flexible conformation than E.coli AspAT.A flexible active site in the rigid T.thermophilus AspAT molecule may explain its high activity even at room temperature.
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Research Products
(8 results)
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[Publications] Kawaguchi, S., Nobe, Y., Yasuoka, J., Wakamiya, T., Kusumoto, S., and Kuramitsu, S.: "Enzyme Flexibility : New Concept in Recognition of Hydrophobic Substrates" J.Biochem.122(No.1). 55-63 (1997)
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「研究成果報告書概要(欧文)」より
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[Publications] Nobe, Y., Kawaguchi, S., Ura, H., Nakai, T., Hirotsu, K., Kato, R., and Kuramitsu, S.: "The Novel Substrate Recognition Mechanism Utilized in Aspartate Aminotransferase of Extreme Thermophile, Thermus thermophilus HB8" J.Biol.Chem.273(No, 45). 29554-29564 (1998)
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「研究成果報告書概要(欧文)」より
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[Publications] Nakai, T., Okada, K., Akutsu, S., Miyahara, I., Kawaguchi, S., Kato, R., Kuramitsu, S., and Hirotsu, K.: "Structure of Thermus thermophilus HB8 Aspartate Aminotransferase and Its Complex with Maleate" Biochemistry. 38(No.8). 2413-2424 (1999)
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「研究成果報告書概要(欧文)」より
