Isolation and characterization of cDNA for human 120 kDa mitochondrial 2,4-dienoyl-coenzyme A reductase.
2,4-Dienoyl-CoA reductase (EC 1.3.1.34) participates in beta-oxidation of (poly)unsaturated enoyl-CoAs and it appears in mammalian mitochondria as two isoforms with molecular masses of 120 and 60 kDa [Hakkola and Hiltunen (1993) Eur. J. Biochem. 215, 199-204]. The 120 kDa isomer is a homotetrameric enzyme, and here we report cDNA cloning of its subunit from human. cDNA clones were isolated by reverse transcriptase-PCR from a fibrosarcoma cell line and by screening from a human liver lambda gt11 cDNA library. The 1128 bp clone contained an open reading frame of 1008 bp encoding a polypeptide of 335 amino acid residues with a predicted molecular mass of 36066 Da. This polypeptide represents the immature monomer of the 120 kDa enzyme, and it contains a predicted N-terminal mitochondrial targeting signal. The amino acid (nucleotide) sequence of human 2,4-dienoyl-CoA reductase shows 82.7% (81.7%) similarity (identity) to the corresponding sequence from the rat. Northern-blot analysis gave a single mRNA species of 1.2 kb in several human tissues, the amounts present in the tissues tested ranking as follows: heart approximately liver approximately pancreas > kidney >> skeletal muscle approximately lung. Immunoblotting of human and rat liver samples with an antibody to the subunit of the rat 120 kDa isoform indicates that the mature human enzyme is larger than its counterpart in the rat. The comparison of amino acid sequences for rat and human enzymes proposes that the difference in the size is 10 amino acid residues. The results show that the rat and human reductases are similar in many characteristics and that the reductase is expressed in human tissues capable of beta-oxidation of fatty acids.
Koivuranta
K T
KT
Biocenter Oulu, Finland.
Hakkola
E H
EH
Hiltunen
J K
JK
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics* / Q000302:isolation & purification*; D044943:Fatty Acid Desaturases / Q000235:genetics*; D015698:Genomic Library; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D008297:Male; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D044925:Oxidoreductases Acting on CH-CH Group Donors; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D012474:Salmon; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D007527:Isoenzymes; D012333:RNA, Messenger; D044943:Fatty Acid Desaturases; D044925:Oxidoreductases Acting on CH-CH Group Donors; C100153:2,4-dienoyl-CoA reductase