Purification and partial characterization of bovine kidney aldehyde dehydrogenase able to oxidize retinal to retinoic acid.
A NAD-dependent enzyme that catalyzes the oxidation of retinal to retinoic acid has been purified to homogeneity from bovine kidney. The procedures used in the purification included ion-exchange chromatography on DEAE-Sepharose, affinity chromatography on Affi-gel blue and chromatography on a Mono-Q anion-exchange column. On the Mono-Q column, the enzyme aldehyde dehydrogenase (ALDH) resolved into two activity peaks designated as ALDH1 and ALDH2. The enzymes ALDH1 and ALDH2 were purified about 114- and 65-fold, respectively. Gel filtration chromatography of the partially purified native enzyme on Sephacryl S-200 HR exhibited a molecular mass of about 108 kDa. Electrophoresis of the purified enzymes under nondenaturing conditions showed a single protein band. However, sodium dodecyl sulfate--polyacrylamide gel electrophorsis indicated three protein bands in the 55, 30, and 22 kDa molecular mass regions. Both enzymes exhibited a broad substrate specificity oxidizing a wide variety of aliphatic and aromatic aldehydes. The ALDH1 enzyme had a pI of 7.45 and exhibited a low Km (6.37 microM) for retinal, while the ALDH2 enzyme was found to have very low Km for acetaldehyde (0.98 microM). Based on its kinetic properties, it is suggested that the ALDH1 enzyme may be the primary enzyme for oxidizing retinal to retinoic acid in bovine kidney.
Bhat
P V
PV
Hôtel-Dieu de Montréal, Department of Medicine, University of Montréal, Canada.
Poissant
L
L
Wang
X L
XL
D000444:Aldehyde Dehydrogenase / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity; D003600:Cytosol / Q000201:enzymology; D007525:Isoelectric Focusing; D007668:Kidney / Q000201:enzymology* / Q000378:metabolism*; D010084:Oxidation-Reduction; D012172:Retinaldehyde / Q000378:metabolism*; D014212:Tretinoin / Q000378:metabolism*
D014212:Tretinoin; D000444:Aldehyde Dehydrogenase; D012172:Retinaldehyde