Henan Provincial Engineering Laboratory of Insect Bio-Reactor and Henan Key Laboratory of Ecological Security for Water Source Region of Mid-Line of South-to-North, Nanyang Normal University, 1638 Wolong Road, Nanyang, Henan, 473061, People's Republic of China.
State Key Laboratory of Automotive Biofuel Technology, 1 Tianguan Avenue, Nanyang, Henan, 473000, People's Republic of China.
Enzyme Microb Technol. 2020 May;136:109513. doi: 10.1016/j.enzmictec.2020.109513. Epub 2020 Jan 18.
L-Phenylglycine (L-PHG) is a member of unnatural amino acids, and becoming more and more important as intermediate for pharmaceuticals, food additives and agrochemicals. However, the existing synthetic methods for L-PHG mainly rely on toxic cyanide chemistry and multistep processes. To provide green, safe and high enantioselective alternatives, we envisaged cascade biocatalysis for the one-pot synthesis of L-PHG from racemic mandelic acid. A engineered E. coli strain was established to co-express mandelate racemase, D-mandelate dehydrogenase and L-leucine dehydrogenase and catalyze a 3-step reaction in one pot, enantioselectively transforming racemic mandelic acid to give L-PHG (e.e. >99 %). After the conditions for biosynthesis of L-PHG optimized by response surface methodology, the yield and space-time yield of L-PHG can reach 87.89 % and 79.70 g·L·d, which was obviously improved. The high-yielding and enantioselective synthetic methods use cheap and green reagents, and E. coli whole-cell catalysts, thus providing green and useful alternative methods for manufacturing L-PHG.
L-苯甘氨酸（L-PHG）是一种非天然氨基酸，作为药物、食品添加剂和农用化学品的中间体，其重要性日益增加。然而，现有的 L-PHG 合成方法主要依赖于有毒的氰化物化学和多步反应。为了提供绿色、安全和高对映选择性的替代方法，我们设想通过级联生物催化，从外消旋扁桃酸一锅法合成 L-PHG。构建了一株能够共表达扁桃酸消旋酶、D-扁桃酸脱氢酶和 L-亮氨酸脱氢酶的工程大肠杆菌菌株，并在一锅反应中催化 3 步反应，对外消旋扁桃酸进行对映选择性转化，得到 L-PHG（ee 值>99%）。通过响应面法优化了 L-PHG 生物合成的条件后，L-PHG 的产率和时空产率分别达到 87.89%和 79.70 g·L·d，明显提高。该高产率和对映选择性的合成方法使用了廉价且绿色的试剂和大肠杆菌全细胞催化剂，为 L-PHG 的制造提供了绿色有用的替代方法。
