| Reversible and nonoxidative gamma-resorcylic acid decarboxylase: characterization and gene cloning of a novel enzyme catalyzing carboxylation of resorcinol, 1,3-dihydroxybenzene, from Rhizobium radiobacter. | |
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MedLine Citation:
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PMID: 15474471 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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We found a gamma-resorcylic acid (gamma-RA, 2,6-dihydroxybenzoic acid) decarboxylase, as a novel enzyme applicable to carboxylation of resorcinol (RE, 1,3-dihydroxybenzene) to form gamma-RA, in a bacterial strain Rhizobium radiobacter WU-0108 isolated through the screening of gamma-RA degrading microorganisms. The activities for carboxylation of RE and decarboxylation of gamma-RA were detected in the cell-free extracts of R. radiobacter WU-0108 grown aerobically with gamma-RA. The enzyme, gamma-RA decarboxylase, was purified to homogeneity on SDS-PAGE through the steps of one ion-exchange chromatography and two kinds of hydrophobic chromatography. The molecular weight of the enzyme was estimated to be 130 kDa by gel-filtration, and that of the subunit was determined to be 34 kDa by SDS-PAGE, suggesting that the enzyme is a homotetrameric structure. The enzyme catalyzed the decarboxylation of gamma-RA, but not alpha-RA or beta-RA. Without addition of any cofactors, the enzyme catalyzed the regio-selective carboxylation of RE to form gamma-RA, without formation of alpha-RA and beta-RA, and of catechol to 2,3-dihydroxybenzoic acid. In the presence of oxygen, this gamma-RA decarboxylase showed no decrease in both of the activities as for decarboxylation of gamma-RA and carboxylation of RE, different from other decarboxylases reported so far. The gene, rdc, encoding the gamma-RA decarboxylase was cloned into Escherichia coli, sequenced, and subjected to over-expression. The deduced amino acid sequence of the rdc gene consists of 327 amino acid residues corresponding to 34 kDa protein, and shows 42% and 30% identity to those of a 2,3-dihydroxybenzoic acid decarboxylase from Aspergillus niger and a 5- carboxyvanillate decarboxylase from Sphingomonas paucimobilis SYK-6. A site-directed mutagenesis study revealed the two histidine residues at positions of 164 and 218 in Rdc to be essential for the catalytic activities of decarboxylation of gamma-RA and carboxylation of RE. |
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Authors:
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Yoshitaka Ishii; Yoshiki Narimatsu; Yuichiro Iwasaki; Naoki Arai; Kuniki Kino; Kohtaro Kirimura |
Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Biochemical and biophysical research communications Volume: 324 ISSN: 0006-291X ISO Abbreviation: Biochem. Biophys. Res. Commun. Publication Date: 2004 Nov |
Date Detail:
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Created Date: 2004-10-11 Completed Date: 2004-12-20 Revised Date: 2006-11-15 |
Medline Journal Info:
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Nlm Unique ID: 0372516 Medline TA: Biochem Biophys Res Commun Country: United States |
Other Details:
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Languages: eng Pagination: 611-20 Citation Subset: IM |
Affiliation:
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Department of Applied Chemistry, School of Science and Engineering, Waseda University, 3-4-1 Ohkubo, Shinjuku-ku, Tokyo 169-8555, Japan. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Aspergillus niger
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metabolism Binding Sites Carbon-Carbon Ligases / chemistry Carboxy-Lyases / chemistry* Carboxylic Acids / chemistry Catalysis Catechols / chemistry Cell-Free System Chromatography, High Pressure Liquid Chromatography, Ion Exchange Chromatography, Thin Layer Cloning, Molecular Cysteine / chemistry DNA / chemistry Dimerization Dose-Response Relationship, Drug Electrophoresis, Polyacrylamide Gel Escherichia coli / metabolism Histidine / chemistry Hydrogen-Ion Concentration Hydroxybenzoic Acids / chemistry* Kinetics Models, Chemical Mutagenesis, Site-Directed Oxygen / chemistry, metabolism* Resorcinols / chemistry* Rhizobium radiobacter / enzymology* Sphingomonas / metabolism Temperature Tretinoin / chemistry |
| Chemical | |
Reg. No./Substance:
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0/Carboxylic Acids; 0/Catechols; 0/Hydroxybenzoic Acids; 0/Resorcinols; 120-80-9/catechol; 302-79-4/Tretinoin; 303-07-1/gamma-resorcylic acid; 52-90-4/Cysteine; 71-00-1/Histidine; 7782-44-7/Oxygen; 9007-49-2/DNA; EC 4.1.1.-/5-carboxyvanillate decarboxylase; EC 4.1.1.-/Carboxy-Lyases; EC 4.1.1.-/gamma-resorcylic acid decarboxylase, Rhizobium radiobacter; EC 6.4.-/Carbon-Carbon Ligases |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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