三酶级联催化L-苏氨酸生产L-2-氨基丁酸

Production of L-2-aminobutyric acid from L-threonine using a trienzyme cascade

摘要:

L-2-氨基丁酸 (L-ABA) 是一种重要的化工原材料和手性医药中间体，为了实现L-ABA的高效生产，本研究在大肠杆菌Escherichia coli BL21 (DE3) 中分别表达大肠杆菌来源的苏氨酸脱氨酶 (Threonine deaminase，TD)、苏云金芽孢杆菌来源的亮氨酸脱氢酶 (Leucine dehydrogenase，LDH) 和博伊丁假丝酵母来源的甲酸脱氢酶 (Formate dehydrogenase，FDH)，构建体外级联酶催化反应实现L-苏氨酸向L-ABA的转化，体系中TD、LDH和FDH添加最适比例为1∶1∶0.2。为了简化生产工艺，将3种酶在一株菌E. coli 3FT+L中共表达并实现上述配比，在30 L发酵罐中用E. coli 3FT+L全细胞转化12 h，L-ABA的产量为68.5 g/L，底物L-苏氨酸的摩尔转化率达到99.0%。该工艺路线绿色高效，为未来大规模生产L-ABA提供借鉴。

Abstract:

L-2-aminobutyric acid (L-ABA) is an important chemical raw material and chiral pharmaceutical intermediate. The aim of this study was to develop an efficient method for L-ABA production from L-threonine using a trienzyme cascade route with Threonine deaminase (TD) from Escherichia. coli, Leucine dehydrogenase (LDH) from Bacillus thuringiensis and Formate dehydrogenase (FDH) from Candida boidinii. In order to simplify the production process, the activity ratio of TD, LDH and FDH was 1:1:0.2 after combining different activity ratios in the system in vitro. The above ratio was achieved in the recombinant strain E. coli 3FT+L. Moreover, the transformation conditions were optimized. Finally, we achieved L-ABA production of 68.5 g/L with a conversion rate of 99.0% for 12 h in a 30-L bioreactor by whole-cell catalyst. The environmentally safe and efficient process route represents a promising strategy for large-scale L-ABA production in the future.