Relationship between enoyl-CoA hydratase and a peroxisomal bifunctional enzyme, enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase, from an n-alkane-utilizing yeast, Candida tropicalis.

Abstract

A protein exhibiting only enoyl-CoA hydratase (EC 4.2.1.17) activity was purified from an n-alkane-grown yeast, Candida tropicalis. This enzyme had a homotetrameric form composed of subunits with a molecular mass of 36kDa. On the other hand, a bifunctional enzyme exhibiting enoyl-CoA hydratase and 3-hydroxyacyl-CoA dehydrogenase (EC 1.1.1.35) activities was obtained from the same yeast cells when purified in the presence of protease inhibitors, phenylmethylsulfonyl fluoride, antipain and chymostatin. The enzyme had a molecular mass of 105 kDa and was a monomeric form. Limited proteolysis of the bifunctional enzyme with α-chymotrypsin yielded a peptide mixture containing a 36 kDa fragment, the mixture showing about 76% of the original enoyl-CoA hydratase activity but no 3-hydroxyacyl-CoA dehydrogenase activity. Comparison of the peptide maps of the purified enoyl-CoA hydratase and the 36 kDa fragment obtained from the bifunctional enzyme showed the similarity of these proteins. These results strongly suggest that the domain of enoyl-CoA hydratase is separable from the bifunctional enzyme through the action of a certain protease.