Purification and characterization of maize leaf acetyl-coenzyme A carboxylase.
Maize leaf acetyl-CoA carboxylase was purified from whole tissue homogenates by precipitation with polyethylene glycol and ammonium sulfate, and gel filtration. Recoveries were approximately 5% with 100-fold increases in specific activity. The molecular weight of the native enzyme is estimated at 500,000 from the elution volume of a calibrated Ultrogel AcA 22 column. Electrophoresis in polyacrylamide gel containing 1% sodium dodecyl sulfate revealed a single subunit of Mr 60,000-61,000. Investigation of the kinetic properties of the purified enzyme indicates that Mg X ATP is the active substrate, with free ATP inhibiting and Mg2+ activating the enzyme. Km's for acetyl-CoA and HCO3- are about 0.1 and 2 mM, respectively. ADP inhibition is competitive with respect to ATP, but uncompetitive with respect to acetyl-CoA. The observed responses of purified acetyl-CoA carboxylase to changes in pH, and in concentrations of Mg2+, ATP, and ADP, and the reported changes in the chloroplastic concentrations of these effectors during light-dark transitions of chloroplasts are consistent with increased acetyl-CoA carboxylase activity upon illumination of chloroplasts.
D000103:Acetyl-CoA Carboxylase / Q000302:isolation & purification / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D004789:Enzyme Activation; D007700:Kinetics; D008025:Ligases / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology; D015636:Magnesium Chloride; D008970:Molecular Weight; D010944:Plants / Q000201:enzymology*; D013379:Substrate Specificity; D003313:Zea mays
D015636:Magnesium Chloride; D000255:Adenosine Triphosphate; D008025:Ligases; D000103:Acetyl-CoA Carboxylase; D008274:Magnesium