cis-1,2-Dihydro-1,2-naphthalenediol (DHND) is a valuable molecule employed for the pharmaceutical synthesis of bioactive compounds, such as bicyclic conduritol analogues. Enantiopure (+)-(1R,2S)-DHND (>98 % ee) is easily biosynthesized through the dearomatizing dihydroxylation of naphthalene, catalyzed by toluene dioxygenase (TDO) from Pseudomonas putida F1. However, the opposite enantiomer (-)-(1S,2R)-DHND could not be directly accessed, neither by chemical synthesis nor via biocatalytic approaches. Herein, we report a one-step biosynthesis of the opposite enantiomer (-)-(1S,2R)-DHND in a recombinant TDO E. coli BW25113 platform. We based on a semi-rational approach to generate a set of TDO variants, targeting exclusively the hotspot position F366, in order to enable an enantiomeric switch in the generated product. Eight out of nine single point variants were active and showed not only an alteration in enantioselectivity, but also generated an enantiomeric excess of the pursued product. Variant TDO_F366V outperformed above the rest of the set, enabling the synthesis of (-)-(1S,2R)-DHND not only with an excellent enantiomeric excess of 90 %, but also with an advantageous product formation. A comparative semi-preparative biosynthesis yielded, 287 mg of (+)-(1R,2S)-DHND (>98 % ee) and 101 mg of (-)-(1S,2R)-DHND (90 % ee), when performed in a total volume of 100 mL with TDO wild-type and TDO_F366V resting cells, respectively.

cis -1,2-Dihydro-1,2-naphthalenediol (DHND) 是一种有价值的分子，用于生物活性化合物的药物合成，例如双环 conduritol 类似物。Enantiopure (+)-(1 R ,2 S )-DHND (>98% ee ) 很容易通过萘的脱芳构化二羟基化进行生物合成，由来自恶臭假单胞菌F1 的甲苯双加氧酶 (TDO) 催化。然而，相反的对映异构体 (-)-(1 S ,2 R )-DHND 无法直接获得，无论是通过化学合成还是通过生物催化方法。在此，我们报告了相反对映异构体 (-)-(1 S ,2 R)-DHND 在重组 TDO大肠杆菌BW25113 平台中。我们基于半合理的方法来生成一组 TDO 变体，专门针对热点位置 F366，以便在生成的产品中启用对映体转换。九个单点变体中有八个是活跃的，不仅显示出对映选择性的改变，而且还产生了所追求产品的对映体过量。变体 TDO _{F366V 的}表现优于该组的其余部分，使 (-)-(1 S ,2 R )-DHND 的合成不仅具有 90% 的优异对映体过量，而且具有有利的产物形成。比较半制备型生物合成产生了 287 mg (+)-(1 R ,2 S)-DHND (>98 % ee ) 和 101 mg (-)-(1 S ,2 R )-DHND (90 % ee )，总体积为 100 mL，TDO 野生型和 TDO _F366V静息细胞，分别。