1996 Fiscal Year Final Research Report Summary
Elucidation of the reaction mechanism of a PCB-degrading enzyme BphyC based on three-dimensional structural information.
| Project/Area Number |
07458251
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Structural biochemistry
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| Research Institution | Nagaoka University of Technology |
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Principal Investigator |
MITSUI Yukio Department of BioEngineering Nagaoka University of Technology, Professor, 工学部, 教授 (40012637)
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| Co-Investigator(Kenkyū-buntansha) |
SENDA Toshiya Department of BioEngineering Nagaoka University of Technology, Instructor, 工学部, 助手 (30272868)
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| Project Period (FY) |
1995 – 1996
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| Keywords | PCB / PCB-degrading enzyme / oxygenase / dioxygenase / catechol ring cleavage / X-ray analysis / protein crystallography / oligomeric enzyme. |
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| Research Abstract |
The so-called BphC enzyme is a dioxygenase which cleaves the catechol ring moiety situated in biphenyl and its derivatives. The cleaving point is located adjacent to the two neighboring hydroxyl groups in a catechol moiety, thus the enzyme works in an extradiol fashion. This kind of enzyme performs such reactions making use of molecular oxygen and incorporates both the atomic oxygen atoms under the catalytic influence of Fe (2+) or ferrous ion.
The present investigators solved the three-dimensional structure of a BphC enzyme from Pseudomonas sp. stran KKS102 by X-ray structure analysis (J.Mol. Biol. 255,735-752 (1996)). The structural information coupled with various enzyme kinetic data on wild and mutant forms of the enzyme revealed the following points.
As for the catalytic mechanism :
1) The site of ring cleavage appears to be specifically determined by the fact that the molecular oxygen binding site is fixed relative to the poisition and orientation of the bound substrate molecules.
2) One of the conserved residues, His 194, plays a role of ctalytic base abstracting a hydrogen from the susceptible hydroxyl group.
As for the substrate specificity :
1) The substrate binding site is essentially hydrophobic exhibiting very high complimentarity to the catechool ring moiety of the substrates.
2) The Fe ion situated in the active site is not necessarily essential for substrate binding (but essential for catalysis, of course).
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[Publications] Senda,T., Sugiyama,K., Narita, H., Yamamoto, T., Kimbara,K., Fukuda, M., Sato,M.,the late Yano, K. & Mitsui, Y.: "Three-dimensional structures of free form and two sustrate complexes of an extradiol ring-cleavage type dioxygenase, the BphC enzyme from Pseudomonas sp. strai KKS 102." J. Mol. Biol.255. 735-752 (1996)
Description
「研究成果報告書概要(和文)」より
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[Publications] Kurihara, J., Nonaka,T., Mitsui,Y., Ohgi, K., Irie, M., & Nakamura,K. T.: "Crystal structure of ribonuclease from Rhizopus niveus at 2.0 A resolution." J. Mol. Biol.255. 310-320 (1996)
Description
「研究成果報告書概要(和文)」より
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[Publications] Tanaka, N., Nonaka,T., Nakanishi, M., Deyashiki, Y.,Hara, A. & Mitsui, Y.: "Crystal structure of the ternary complex of mouse lung carbonyl reductase at 1.8 A resolution : The structural origin of distinct coenzyme specificities among the enzymes of the short-chain dehydrogenase/reductase family." Structure. 4. 33-45 (1996)
Description
「研究成果報告書概要(和文)」より
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[Publications] Orita, M., Nishikawa, F., Kohno,T., Senda,T., Mitsui,Y., Endo,Y., Taira, K. & Nishikawa, S.: "High-resolution NMR study of a GdA GA tetranucleotide loop that is an improved substrate for ricin, a cytotoxia plant protein." Nucleic Acid Research. 24. 611-618 (1996)
Description
「研究成果報告書概要(和文)」より
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[Publications] Tanaka, N., Nonaka, T., Tanabe, T., Yoshimoto, T., Tsuru, D. & Mitsui, Y.: "Crystal structures of the binary and ternary complexes of 7α-hydroxy-steroid dehydrogenase from E. coli." Biochemistry. 35. 7715-7724 (1996)
Description
「研究成果報告書概要(和文)」より
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[Publications] Nakanishi, M., Kakumoto, M., Matsuura,K., Deyashiki, Y., Tanaka,N., Nonaka, T., Mitsui, Y. & Hara, A.: "Involvement of two basic residues (Lys-17 and Arg-39) of mouse lung carbonyl reductase in NADP (H)-binding and fatty acid activation : Site-directed mutagenesis and kinetic analyses." J. Biochem. (Tokyo). 120. 257-263 (1996)
Description
「研究成果報告書概要(和文)」より
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[Publications] Nakanishi, M., Natsuura, K., Kaibe,H., Tanaka,N., Nonaka, T., Mitsui, Y. & Hara, A.: "Switch of coenzyme specificity of mouse lung carbonyl reductase by substitution of Threonine 38 with Aspartic acid." J. Biol. Chem.272. 2218-2222 (1997)
Description
「研究成果報告書概要(和文)」より
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-
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[Publications] Senda, T., Sugiyama, K., Narita, H., Yamamoto, T., Kimbara, K., Fukuda, M., Sato, M., the late Yano, K.& Mitsui, Y.: "Three-dimensional structures of free form and two sustrate complexes of an extradiol ring-cleavage type dioxygenase, the BphC enzyme from Pseudomonas sp. strain KKS102." J.Mol. Biol.255. 735-752 (1996)
Description
「研究成果報告書概要(欧文)」より
-
[Publications] Kurihara, J., Nonaka, T., Mitsui, Y., Ohgi, K., Irie, M.& Nakamura, K.T.: "Crystal structure of ribonuclease from Rhizopus niveus at 2.0A resolution." J.Mol. Biol.255. 310-320 (1996)
Description
「研究成果報告書概要(欧文)」より
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[Publications] Tanaka, N., Nonaka, T., Nskanishi, M., Deyashiki, Y., Hara, A.& Mitsui, Y.: "Crystal structure of the ternary comnplex of mouse lung carbonyl reductase at 1.8A resolution : The structural origin of distinct coenzyme specificities among the enzymes of the short-chain dehydrogenase/reductase family." Structure. 4. 33-45 (1996)
Description
「研究成果報告書概要(欧文)」より
-
[Publications] Orita, M., Nishikawa, F., Kohno, T., Senda, T., Mitsui, Y., Endo, Y., Taira, K.& Nishikawa, S.: "High-resolution NMR study of a GdA GA tetranucleotide loop that is an improved substrate for ricin, a cytotoxic plant protein." Nucleic Acid Research. 24. 611-618 (1996)
Description
「研究成果報告書概要(欧文)」より
-
[Publications] Tanaka, N., Nonaka, T., Tanabe, T., Yoshimoto, T., Tsuru, D.& Mistui, Y.: "Crystal structure of the binary and ternary and ternary complexes of 7alpha-hydroxy-steroid dehydrogenase from E.coli." Biochemistry. 35. 7715-7724 (1996)
Description
「研究成果報告書概要(欧文)」より
-
[Publications] Nakanishi, M., Kakumoto, M., Matsuura, K., Deyashiki, Y., Tanaka, N., Nonaka, T., Mitsui, Y.& Hara, A..: "Involvement of two basic residues (Lys-17 and Arg-39) of mouse lung carbonyl reductase in NADP (H) -binding and fatty acid activation : Site-directed mutagenesis and kinetic' analyzes." J.Biochem. (Tokyo). 120. 257-263 (1996)
Description
「研究成果報告書概要(欧文)」より
-
[Publications] Nakanishi, M., Natsuura, K., Kaibe, H., Tanaka, N., Nonaka, T., Mitsui, Y.& Hara, A.: "Switch of coenzyme specificitiy of mouse lung carbonyl reductase by substitution of Threonine 38 with Aspartic acid." J.Biol. Chem.272. 2218-2222 (1997)
Description
「研究成果報告書概要(欧文)」より
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