NADP-specific isocitrate dehydrogenase of Escherichia coli. IV. Purification by chromatography on Affi-Gel Blue.
Affinity chromatography on Affi-Gel Blue has been used to purify the NADP-specific isocitrate dehydrogenase (EC 1.1.1.42) from Escherichia coli. The protocol permits rapid purification of the enzyme in milligram quantities with a yield of 50% and is carried out almost entirely at room temperature. The preparation was judged to be homogeneous by non-denaturing electrophoresis at pH 7.5 and denaturing electrophoresis in the presence of sodium dodecyl sulfate. The subunit molecular weight of 53 000, determined by sodium dodecyl sulfate gel electrophoresis, is in reasonable agreement with the value of 46 900 estimated from the amino acid composition data.
D000596:Amino Acids / Q000032:analysis; D002846:Chromatography, Affinity / Q000379:methods; D004926:Escherichia coli / Q000201:enzymology*; D007521:Isocitrate Dehydrogenase / Q000032:analysis / Q000302:isolation & purification*; D009249:NADP; D013045:Species Specificity; D013379:Substrate Specificity
D000596:Amino Acids; D009249:NADP; D007521:Isocitrate Dehydrogenase