Molecular cloning and characterization of a mitochondrial peroxisome proliferator-induced acyl-CoA thioesterase from rat liver.
We have previously reported the purification and characterization of the peroxisome proliferator-induced very-long-chain acyl-CoA thioesterase (MTE-I) from rat liver mitochondria [L.T. Svensson, S.E. H. Alexson and J.K. Hiltunen (1995) J. Biol. Chem. 270, 12177-12183]. Here we describe the cloning of the corresponding cDNA. One full-length clone was isolated that contained an open reading frame of 1359 bp encoding a polypeptide with a calculated molecular mass of 49707 Da. The deduced amino acid sequence contains a putative mitochondrial leader peptide of 42 residues. Expression of the cDNA in Chinese hamster ovary cells, followed by immunofluorescence, immunoelectron microscopy and Western blot analyses, showed that the product was targeted to mitochondria and processed to a mature protein of 45 kDa, which is similar to the molecular mass of the protein isolated from rat liver mitochondria. The recombinant enzyme showed the same acyl-CoA chain-length specificity as the isolated rat liver enzyme. Sequence analysis showed no similarity to known esterases, but a high degree (approx. 40%) of identity with bile acid-CoA:amino acid N-acyltransferase cloned from human and rat liver. A putative active-site serine motif (Gly-Xaa-Ser-Xaa-Gly) of several carboxylesterases and lipases was identified. Western and Northern blot analyses showed that MTE-I is constitutively expressed in heart and is strongly induced in liver by feeding rats with di(2-ethylhexyl)phthalate, a peroxisome proliferator, suggesting a role for the enzyme in lipid metabolism.
Svensson
L T
LT
Department of Biochemistry, Shinshu University School of Medicine, Nagano, Japan.
Engberg
S T
ST
Aoyama
T
T
Usuda
N
N
Alexson
S E
SE
Hashimoto
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary / Q000302:isolation & purification; D004051:Diethylhexyl Phthalate / Q000494:pharmacology; D004790:Enzyme Induction / Q000187:drug effects; D007527:Isoenzymes / Q000302:isolation & purification; D008297:Male; D008830:Microbodies / Q000201:enzymology*; D008930:Mitochondria, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000187:drug effects / Q000235:genetics; D010170:Palmitoyl-CoA Hydrolase / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000201:enzymology
D018076:DNA, Complementary; D007527:Isoenzymes; D012333:RNA, Messenger; D004051:Diethylhexyl Phthalate; D010170:Palmitoyl-CoA Hydrolase