For improving the enantioselectivity of Phaseolus vulgaris epoxide hydrolase1 (PvEH1) towards p-chlorostyrene oxide (pCSO)

three mutations (PvEH1W102L

PvEH1P137K and PvEH1I151V) were constructed by site-directed mutagenesis. Firstly

catalytic characteristics of the enzymatic hydrolysis of rac-pCSO by the whole cell of E. coli expressing PvEH1W102L

PvEH1P137K or PvEH1I151V were studied respectively. After then

the rac-pCSO initial concentration in the kinetic resolution of rac-pCSO by PvEH1W102L were optimized

in additional the course was monitored to determine the reaction time. At last

the mechanism of PvEH1W102L's improvement in enantioselectivity towards pCSO was analyzed by molecular simulation. The result indicated that

the E value and relative activity of the best mutation PvEH1W102L were 25.5 and 212.6%

which were 11.6 and 2.1 times of PvEH1

respectively. After four-hour reaction

the yield of (R)-pCSO in the kinetic resolution was 45.62% (ees=96.30%)

when the yield of (R)-p-chlorophenyl-1

2-ethanediol was 50.91% (eep=90.26%). Molecular simulation analysis showed that the mutation of the tryptophan (W) at position 102 of PvEH1 to leucine (L) reduced the binding ability of the enzyme to (R)-pCSO

thereby increasing the enantioselectivity of PvEH1W102L to pCSO.