EXPRESSION OF ACETOACETYL-COA THIOLASE ISOZYME GENES OF N-ALKANE-ASSIMILATING YEAST, CANDIDA-TROPICALIS - ISOZYMES IN 2 INTRACELLULAR COMPARTMENTS ARE DERIVED FROM THE SAME GENES

In the n-alkane-assimilating yeast Candida tropicalis, there are two i sozymes of acetoacetyl-CoA thiolase, peroxisomal acetoacetyl-CoA thiol ase (peroxisomal Thiolase I), and cytosolic acetoacetyl-CoA thiolase ( cytosolic Thiolase I). We have previously isolated two genes (CT-T1A a nd CT-T1B) which encode Thiolase I. In order to compare the expressed products of Thiolase I isozyme-encoding genes in C. tropicalis, cytoso lic Thiolase I was first purified from glucose-grown C. tropicalis in which the proliferation of peroxisomes and the expression of peroxisom al Thiolase I were repressed. Cytosolic Thiolase I was virtually ident ical to peroxisomal Thiolase I in molecular mass, kinetic and immunoch emical properties, and primary structure at the N-terminus. Amino acid sequence analysis revealed that cytosolic Thiolase I was the mixture of products of two genes (CT-T1A and CT-T1B), as in the case of the pe roxisomal enzyme. CT-T1A and CT-T1B were expressed independently in th e yeast Saccharomyces cerevisiae and the recombinant proteins were pur ified. Recombinant Thiolase IA and IB exhibited practically identical enzymatic properties to cytosolic and peroxisomal Thiolase Is from C. tropicalis. These results revealed that cytosolic Thiolase I and perox isomal Thiolase I were encoded not by different genes, but by the same genes (CT-T1A and CT-T1B) and are present as a mixture of products ex pressed by both genes, although their subcellular localizations are di fferent.