| Co-Investigator(Kenkyū-buntansha) |
NAGASAWA Toru Gifu University, Faculty of Engineering, Professor, 工学部, 教授 (60115904)
MITSUKURA Koichi Gifu University, Faculty of Engineering, Research Associate, 工学部, 助手 (70324283)
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| Budget Amount *help |
¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 2005: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2004: ¥2,900,000 (Direct Cost: ¥2,900,000)
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| Research Abstract |
Various enzymes catalyzing oxidative or nonoxidative decarboxylation reaction are involved in aerobic or anaerobic degradation of aromatic carboxylic acids in microorganisms. Recently, we found and characterized two kinds of novel decarboxylase, catalyzing the reverse carboxylation reaction efficiently. The findings strongly suggested the universal occurrence of an enzyme group reversibly catalyzing the decarboxylation of aromatic carboxylates. In the present study, we surveyed and characterized other reversible decarboxylases, and then applied them to the synthesis for aromatic hydroxycarboxylic acids.
We found a bacgerium, Pandoraea sp.12B-2, of which whole cells catalyzed not only the decarboxylation of 2,6-dihydroxybenzoate but also the regioselective carboxylation of 1,3-dihydroxybenzene to 2,6-dihydroxybenzoate. The whole cells of the bacterium also catalyzed the regioselective carboxylation of phenol and 1,2-dihydroxybenzene to 4-hydroxybenzoate and 2,3-dihydroxybenzoate, respectively. The molar conversion ratio of the carboxylation reaction depended on the concentration of KHCO_3 in the reaction mixture. About 50% of 1,3-dihydroxybenzene added was converted into 2,6-dihydroxybenzoate in the presence of 3 M KHCO_3. When the efficient production of 2,6-dihydroxybenzoate was optimized, the. productivity of 2,6-dihydroxybenzoate reached to 1.43 M, which was he highest value so far reported. No formation of any other products was observed after the carboxylation reaction.
The genes encoding 2,6-dihydroxybenzoate decarboxylase of Pandoraea sp.12B-2 and Agrobacterium tumefaciens IAM12048 were isolated and cloned. The deduced primary structures of the decarboxylases showed no homology with various decarboxylases and carboxylases reported. Further analysis on primary structures demonstated that reversible decarboxylases were classified into two subgroups, structures, various reversible decarboxylase were classified into two subgroups.
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