Studies on the subunits of Escherichia coli coenzyme A transferase. Reconstitution of an active enzyme.
The alpha and beta subunits of the acetyl-CoA:acetoacetate-CoA transferase were purified by isoelectric focusing of the enzyme in the presence of 6 M urea. The purified beta subunit, in which the active center of the enzyme is located, exhibits low catalytic activity (2% of the specific activity of the native enzyme) which is stimulated 5-6-fold in the presence of an equimolar concentration of alpha subunit. The presence of the substrate,acetoacetyl-CoA, is  required to recover the catalytic activity of the beta subunit and mixtures containing purified alpha and beta subunits. When the enzyme is dissociation in the presence of 6 M urea and the subunits are not fractioned, removal of the urea by dialysis results in the recovery of 88-98% of enzymic activity and the native alpha2beta2 subunit structure. However, analysis of this renatured enzyme by immunochemical techniques shows that the enzyme does not refold to a completely native conformation. This renatured enzyme exhibits an immunological reactivity more closely resembling the isolated alpha subunit. The results indicate that the alpha subunit serves as a structural subunit, or possible a maturation subunit, imposing a conformation on the beta subunit that is catalytically more competent.
D000101:Acetyl-CoA C-Acetyltransferase / Q000302:isolation & purification* / Q000378:metabolism; D000123:Acetyltransferases / Q000302:isolation & purification*; D000596:Amino Acids / Q000032:analysis; D002384:Catalysis; D004926:Escherichia coli / Q000201:enzymology*; D010446:Peptide Fragments
D000596:Amino Acids; D010446:Peptide Fragments; D000123:Acetyltransferases; D000101:Acetyl-CoA C-Acetyltransferase