MOLECULAR-CLONING AND NUCLEOTIDE-SEQUENCE OF THE CDNA FOR RAT PEROXISOMAL ENOYL-COA - HYDRATASE-3-HYDROXYACYL-COA DEHYDROGENASE BIFUNCTIONAL ENZYME

Abstract

The induction mechanism of peroxisomal .beta.-oxidation enzymes and the biogenesis of the organelle, was studied in isolated [complementary] cDNA clones for rat peroxisomal enoyl-CoA: hydratase-3-hydroxyacyl-CoA dehydrogenase bifunctional enzyme. On blotting experiments with liver RNA, the cDNA hybridized to a 3.0-kilobase RNA which was increased 5-7-fold by the administration of di-(2-ethylhexyl)phthalate to rats. Nucleotide sequencing was carried out for 4 cloned cDNA and 1 obtained by a primer extension method. By overlapping these sequences with each other 20 nucleotides of 5''-noncoding, 2166 nucleotides of coding and 910 nucleotides of 3''-noncoding regions were identified. The deduced amino acid sequence of the enzyme is composed of 722 residues, and the composition agrees with that of the protein data. The sequence was confirmed by the amino acid compositions and sequence analyses of some of the tryptic peptides. The MW of the mature enzyme is calculated to be 78,511 from the predicted amino acid sequence. The enzyme has no terminal peptide extension as a signal for translocation into peroxisomes.