Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science & Technology, Tianjin 300457, China; National Engineering Laboratory for Industrial Enzymes and Tianjin Engineering Research Center of Biocatalytic Technology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 32 Xi Qi Dao, Tianjin Airport Economic Area, Tianjin, 300308, China.
National Engineering Laboratory for Industrial Enzymes and Tianjin Engineering Research Center of Biocatalytic Technology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 32 Xi Qi Dao, Tianjin Airport Economic Area, Tianjin, 300308, China.
Enzyme Microb Technol. 2018 Jun;113:37-43. doi: 10.1016/j.enzmictec.2018.02.008. Epub 2018 Feb 21.
Reversible benzoic acid decarboxylases are versatile biocatalysts by taking advantage of both decarboxylation and carboxylation reactions, especially for the biocatalytic Kolbe-Schmitt reaction. In the course of developing a benzoic acid decarboxylase tool-box, a putative benzoic acid decarboxylase gene from Fusarium oxysporum was heterologously over-expressed in Escherichia coli, the recombinant protein was purified and characterized. The purified enzyme exhibited relatively high catalytic efficiencies for the decarboxylation of 2, 3-dihydroxybenzoic acid and carboxylation of catechol (k/K = 2.03 × 10 and 1.88 mM min, respectively), and thus characterized as 2, 3-dihydroxybenzoic acid decarboxylase (2, 3-DHBD_Fo). The enzyme also catalyzed the decarboxylation of various substituted salicylic acids with different groups at varied positions except 5-position and the carboxylation of phenol and the substituted phenols. In a preparative reaction, catechol was carboxylated into 2, 3-dihydroxybenoic acid with 95% conversion by adding dodecyldimethylbenzylammonium chloride into the reaction system, and the product was isolated in 72% yield. These results demonstrate that 2, 3-DHBD_Fo is a valuable addition to the benzoic acid decarboxylase tool-box with potential practical applications.
利用脱羧和羧化反应，可逆的苯甲酸脱羧酶是一种多功能的生物催化剂，特别是对于生物催化的Kolbe-Schmitt 反应。在开发苯甲酸脱羧酶工具包的过程中，从尖孢镰刀菌中异源过表达了一种假定的苯甲酸脱羧酶基因，对重组蛋白进行了纯化和表征。该酶对 2,3-二羟基苯甲酸的脱羧和儿茶酚的羧化表现出相对较高的催化效率（k/K=2.03×10 和 1.88 mM min，分别），因此被鉴定为 2,3-二羟基苯甲酸脱羧酶（2,3-DHBD_Fo）。该酶还催化各种取代的水杨酸的脱羧，取代基位于不同位置（除 5 位），酚和取代酚的羧化。在制备反应中，通过向反应体系中添加十二烷基二甲基苄基氯化铵，将儿茶酚羧化为 2,3-二羟基苯甲酸，转化率为 95%，产物收率为 72%。这些结果表明，2,3-DHBD_Fo 是苯甲酸脱羧酶工具包的有价值的补充，具有潜在的实际应用价值。
