Purification of acetate kinase by affinity chromatography.
A one-step procedure using affinity chromatography has been shown to purify to apparent homogeneity acetate kinase from a commercially available preparation and to partially purify the enzyme from a crude, cell-free extract. Since the gel's capacity for enzyme adsorption is controlled by the thermodynamics of ligand-enzyme interaction, maximization of the adsorption isotherm was attempted. Enzyme adsorption decreased logarithmically with increasing ionic strength but increased with increasing concentration of MgCl2. These competing effects caused the net adsorption of enzyme to increase to a maximum and then to decrease as the MgCl2 concentration was raised. The results allow a significant improvement in affinity column performance and have important implications for scale-up procedures.
D000084:Acetate Kinase / Q000302:isolation & purification* / Q000378:metabolism; D000327:Adsorption; D002846:Chromatography, Affinity / Q000379:methods; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D008274:Magnesium; D009994:Osmolar Concentration; D010770:Phosphotransferases / Q000302:isolation & purification*
D010770:Phosphotransferases; D000084:Acetate Kinase; D008274:Magnesium