LplR, a Repressor Belonging to the TetR Family, Regulates Expression of the l-Pantoyl Lactone Dehydrogenase Gene in Rhodococcus erythropolis

Abstract

ABSTRACTThel-pantoyl lactone (l-PL) dehydrogenase (LPLDH) gene (lpldh) has been cloned fromRhodococcus erythropolisAKU2103, and addition of 1,2-propanediol (1,2-PD) was shown to be required forlpldhexpression in this strain. In this study, based on an exploration of the nucleotide sequence aroundlpldh, a TetR-like regulator gene, which we designatedlplR, was found upstream oflpldh, and three putative open reading frames existed between the two genes. Disruption oflplRled to 22.8 times higherlpldhexpression, even without 1,2-PD induction, than that in wild-typeR. erythropolisAKU2103 without 1,2-PD addition. Introduction of a multicopy vector carryinglplR(multi-lplR) into the wild-type and ΔlplRstrains led to no detectable LPLDH activity even in the presence of 1,2-PD. The results of an electrophoretic mobility shift assay revealed that purified LplR bound to a 6-bp inverted-repeat sequence located in the promoter/operator region of the operon containinglpldh. These results indicated that LplR is a negative regulator inlpldhexpression. Based on the clarification of the expression mechanism oflpldh, recombinant cells showing high LPLDH activity were constructed and used as a catalyst for the conversion ofl-PL to ketopantoyl lactone. Finally, a promising production process ofd-PL fromdl-PL was constructed.