Cloning of a cDNA for a second retinol dehydrogenase type II. Expression of its mRNA relative to type I.
A retinol dehydrogenase, RoDH(1), which recognizes holo-cellular retinol-binding protein (CRBP) as substrate, has been cloned, expressed, and identified as a short-chain dehydrogenase/reductase (Chai, X., Boerman, M. H. E. M., Zhai, Y., and Napoli, J. L. (1995) J. Biol. Chem. 270, 3900-3904). This work reports the cloning and expression of a cDNA encoding a RoDH isozyme, RoDH(II). The predicted amino acid sequence verifies RoDH(II) as a short-chain dehydrogenase/reductase, 82% identical with RoDH(I). RoDH(II) recognized the physiological form of retinol as substrate, CRBP, with a Km of 2 mM. Similar to microsomal RoDH and RoDH(I), RoDH(II) had higher activity with NADP rather than NAD, was stimulated by ethanol and phosphatidyl choline, was not inhibited by the medium-chain alcohol dehydrogenase inhibitor 4-methylpyrazole, but was inhibited by phenylarsine oxide and the short-chain dehydrogenase/reductase inhibitor carbenoxolone. Northern blot analysis detected RoDH(I) and RoDH(II) mRNA only in rat liver, but RNase protection assays revealed RoDH(I) and RoHD(II) mRNA in kidney, lung, testis, and brain. These data indicate that short-chain dehydrogenases/reductase isozymes expressed tissue-distinctively catalyze the first step of retinoic acid biogenesis from the physiologically most abundant substrate, CRBP.
D000429:Alcohol Oxidoreductases / Q000096:biosynthesis* / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D000072467:Cytochrome P450 Family 2; D017931:DNA Primers; D018076:DNA, Complementary; D015870:Gene Expression; D007527:Isoenzymes / Q000096:biosynthesis* / Q000737:chemistry; D008099:Liver / Q000201:enzymology; D008297:Male; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009928:Organ Specificity; D017433:Protein Structure, Secondary; D012333:RNA, Messenger / Q000096:biosynthesis*; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D012333:RNA, Messenger; D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C022520:retinol dehydrogenase; C504568:Cyp2d1 protein, rat; D000072467:Cytochrome P450 Family 2