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Functionalization of aromatic compounds by decarboxylases catalyzing regioselective carboxylation

脱羧酶催化区域选择性羧化对芳香族化合物进行官能化

基本信息

批准号：
16580056
负责人：
YOSHIDA Toyokazu
金额：
$ 2.43万
依托单位：
Gifu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

项目摘要

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.

催化氧化或非氧化脱羧反应的各种酶参与微生物中芳香族羧酸的需氧或厌氧降解。最近，我们发现并表征了两种新型脱羧酶，可有效催化逆羧化反应。这些发现强烈表明普遍存在可逆催化芳香族羧酸盐脱羧的酶基团。在本研究中，我们对其他可逆脱羧酶进行了调查和表征，然后将它们应用于芳香族羟基羧酸的合成。我们发现了一种细菌，Pandoraea sp.12B-2，其全细胞不仅催化2,6-二羟基苯甲酸的脱羧，而且还催化2,6-二羟基苯甲酸的区域选择性羧化。 1,3-二羟基苯转化为2,6-二羟基苯甲酸酯。该细菌的全细胞还催化苯酚和1,2-二羟基苯分别区域选择性羧化为4-羟基苯甲酸酯和2,3-二羟基苯甲酸酯。羧化反应的摩尔转化率取决于反应混合物中KHCO_3的浓度。添加的约50%的1,3-二羟基苯在3M KHCO_3存在下转化为2,6-二羟基苯甲酸酯。当优化 2,6-二羟基苯甲酸酯的高效生产时，。 2,6-二羟基苯甲酸酯的产率达到1.43 M，这是迄今为止报道的最高值。羧化反应后未观察到任何其他产物的形成。分离并克隆了Pandoraea sp.12B-2和根癌农杆菌IAM12048的2,6-二羟基苯甲酸脱羧酶基因。推导的脱羧酶的一级结构与报道的各种脱羧酶和羧化酶没有同源性。进一步对一级结构的分析表明，可逆性脱羧酶分为两个亚类，结构上，各种可逆性脱羧酶分为两个亚类。