Identification and characterization of all-trans-retinol dehydrogenase from photoreceptor outer segments, the visual cycle enzyme that reduces all-trans-retinal to all-trans-retinol.
Retinol dehydrogenase (RDH), the enzyme that catalyzes the reduction of all-trans-retinal to all-trans-retinol within the photoreceptor outer segment, was the first visual cycle enzymatic activity to be identified. Previous work has shown that this enzyme utilizes NADPH, shows a marked preference for all-trans-retinal over 11-cis-retinal, and is tightly associated with the outer segment membrane. This paper reports the identification of a novel member of the short chain dehydrogenase/reductase family, photoreceptor RDH (prRDH), using subtraction and normalization of retina cDNA, high throughput sequencing, and data base homology searches to detect retina-specific genes. Bovine and human prRDH are highly homologous and are most closely related to 17-beta-hydroxysteroid dehydrogenase 1. The enzymatic properties of recombinant bovine prRDH closely match those previously reported for RDH activity in crude bovine rod outer segment preparations. In situ hybridization and RNA blotting show that the PRRDH gene is expressed specifically in photoreceptor cells, and protein blotting and immunocytochemistry show that prRDH localizes exclusively to both rod and cone outer segments and that prRDH is tightly associated with outer segment membranes. Taken together, these data indicate that prRDH is the enzyme responsible for the reduction of all-trans-retinal to all-trans-retinol within the photoreceptor outer segment.
Rattner
A
A
Department of Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Smallwood
P M
PM
Nathans
J
J
D000429:Alcohol Oxidoreductases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D006801:Humans; D008969:Molecular Sequence Data; D009243:NAD / Q000378:metabolism; D012164:Retinal Diseases / Q000209:etiology; D012172:Retinaldehyde / Q000378:metabolism*; D012374:Rod Cell Outer Segment / Q000201:enzymology*; D014801:Vitamin A / Q000378:metabolism*
D009243:NAD; D014801:Vitamin A; D000429:Alcohol Oxidoreductases; C113992:trans-retinol dehydrogenase; D012172:Retinaldehyde