Characterization of Aldose Reductase and Aldehyde Reductase from the Medulla of Rat Kidney

Abstract

Aldos reductase and aldehyde reductase from the medulla of the rat kidney have been purified to homogeneity by using affinity chromatography, gel filtration and chromatofocusing. The molecular weights of aldose reductase and aldehyde reductase by sodium dodecyl sulfate (SDS)-polyacylamide gel electrophoresis were found to be 37000 and 39000, rspectively. The isoelectric points of aldose reductase and aldehyde reductase were found to be 5.4 and 6.2 by chromatofocusing, respectively. The major differences of amino acid compositinos between both enzymes were fonud in serine, alanine and aspartic acid. Substrate specificity studies showed that aldose reductase utilizad aldo-sugars such ans D-glucose and D-galactose, but aldehyde reductase did not use them. The K_m values of aldose reductase for various substrates were lower than those of aldehyde reductase. Aldose reductase utilized both reduced nicotinamide adenine dinucleotide phosphate (NADPH) and reduced nicotinamide adenine dinucleotide (NADH) as coenzymes, whereas aldehyde reductase utilized only NADPH. The presence of the sulfate ion resulted in a dramatic activation of aldose reductase whereas it did not affect aldehyde reductase activity. These enzymes ewre strongly inhibited by the known aldose reductase inhibitors. However, aldose reductase was more susceptible than aldehyde reductase to inhibition by the aldose reductase inhibitors.