Stable preparation of aldose reductase isoenzymes from human placenta.
An efficient, large-scale purification has been achieved for two aldose reductase isoenzymes from human placenta in stable form. The procedure included ammonium sulfate fractionation (45-75%), followed by chromatographies on Matrex Red A, DE-52 cellulose, and Matrex Orange A. The preparations were stable for at least 3 months at 3 degrees C. IC50 values toward sorbinil were similar to those reported for crude or partially purified enzymes, indicating that they retained native structures during the purification steps. The molecular weights of purified GAR1 and GAR2, named according to their order of elution with a salt gradient from a Matrex Red A column, were 36,600 and 40,300, respectively. Kinetic studies indicate that GAR1 belongs to an aldose reductase (a low-Km form) and GAR2 to an aldehyde reductase (a high-Km form). GAR2, an aldehyde reductase, was also active in the reduction of D-glucose, with an apparent Km comparable to that of GAR1 but with a Vmax only 14% that of GAR1.
Fujii
Y
Y
Department of Chemistry, Fukui Medical School, Japan.
Zhao
F X
FX
Fu
S C
SC
Nakai
N
N
Lai
C Y
CY
D000449:Aldehyde Reductase / Q000302:isolation & purification* / Q000378:metabolism; D000645:Ammonium Sulfate; D005591:Chemical Fractionation; D002845:Chromatography / Q000379:methods; D004795:Enzyme Stability; D005260:Female; D006801:Humans; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D013379:Substrate Specificity; D013401:Sugar Alcohol Dehydrogenases / Q000302:isolation & purification / Q000378:metabolism
D007527:Isoenzymes; D013401:Sugar Alcohol Dehydrogenases; C028041:glyceraldehyde reductase; D000449:Aldehyde Reductase; D000645:Ammonium Sulfate