Abstract

Chiral dimethyl 2-methylsuccinate (1) is a very important building block for the manufacturing of many active pharmaceutical ingredients and fine chemicals. The asymmetric reduction of C=C double bond of dimethyl citraconate (2), dimethyl mesaconate (3) or dimethyl itaconate (4) by ene-reductases (ERs) represents an attractive straightforward approach, but lack of high-performance ERs, especially (S)-selective ones, has limited implementing this method to prepare the optically pure dimethyl 2-methylsuccinate. Herein, three ERs (Bac-OYE1 fromBacillussp.,SeER fromSaccharomyces eubayanusandAfER fromAspergillus flavus) with high substrate tolerance and stereoselectivity towards2,3and4have been identified. Up to 500 mM of3was converted to (S)-dimethyl 2-methylsuccinate ((S)-1) bySeER in high yields (80%) and enantioselectivity (98%ee), and 700 mM of2and 400 mM of4were converted to (R)-1by Bac-OYE1 andAfER, respectively, in high yields (86% and 77%) with excellent enantioselectivity (99%ee). The reductions of diethyl citraconate (5), diethyl mesaconate (6) and diethyl itaconate (7) were also tested with the three ERs. Although up to 500 mM of5was completely converted to (R)-diethyl 2-methylsuccinate ((R)-8) by Bac-OYE1 with excellent enantioselectivity (99%ee), the alcohol moiety of the esters had a great effect on the activity and enantioselectivity of ERs. This work provides an efficient methodology for the enantiocomplementary production of optically pure dimethyl 2-methylsuccinate from dimethyl itaconate and its isomers at high titer.Keywords:dimethyl citraconate;dimethyl itaconate;dimethyl mesaconate;dimethyl 2-methylsuccinate;enantioselective enzymatic reduction;ene-reducase