Substrate specificity and stereospecificity of limonene-1,2-epoxide hydrolase from Rhodococcus erythropolis DCL14; an enzyme showing sequential and enantioconvergent substrate conversion

Limonene-1,2-epoxide hydrolase (LEH) from Rhodococcus erythropolis DCL14, a n enzyme involved in the limonene degradation pathway of this microlorganis m, has a narrow substrate specificity. Of the compounds tested, the natural substrate, limonene-1,2-epoxide, and several alicyclic and 2-methyl-1,2-ep oxides (e.g. 1-methylcyclohexene oxide and indene oxide), were substrates f or the enzyme. When LEH was incubated with a diastereomeric mixture of limo nene-1,2-epoxide, the sequential hydrolysis of first the (1R,2S)- and then the (1S,2R)-isomer was observed. The hydrolysis of (4R)- and (4S)-limonene- 1,2-epoxide resulted in, respectively, (1S,2S,4R)- and (1R,2R,4S)-limonene- 1,2-diol as the sole product with a diastereomeric excess of over 98%. With all other substrates, LEH showed moderate to low enantioselectivities (E r atios between 34 and 3).