Separation of the isoenzymes of glyoxalase I from human red blood cells by electrophoresis and isoelectric focusing on polyacrylamide gel and by ion exchange chromatography.
Methods have been devised for the separation of the isoenzymes of glyoxalase I(S-lactoylglutathione methylglyoxal-lyase (isomerizing), EC 4.4.1.5) from human red blood cells by electrophoresis and electrofocusing on polyacrylamide gel slabs. Three different staining methods were used for the location of the enzyme. Three electrophoretic phenotypes of the enzyme were resolved, the fast and slow types with one band and the intermediate type with three glyoxalase I activity bands. In gel electrofocusing (pH gradient 3.5-9.5) two glyoxalase I activity bands were found for all electrophoretic types. In electrofocusing on gel with a narrow pH gradient, at least four separate enzyme components were resolved for the fast and slow electrophoretic types and at least six components for the intermediate type. The phenotypes could be distinguished correspondingly to the electrophoretic results. Preparative separation of the isoenzymes were achieved by ion exchange chromatography on DEAE-Sephacel but gel chromatography on Sephadex G-100 gave the same elution volume for all enzyme phenotypes. This corresponds to an apparent molecular weight of about 47 000.
D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000097:blood*; D007791:Lactoylglutathione Lyase / Q000097:blood*; D008190:Lyases / Q000097:blood*; D008722:Methods; D013194:Staining and Labeling
D007527:Isoenzymes; D008190:Lyases; D007791:Lactoylglutathione Lyase