Evidence for a protein regulator from rat liver which activates acetyl-CoA carboxylase.
1. A regulator of acetyl-CoA carboxylase has been identified in high-speed supernatant fractions from rat liver. The regulator was found to activate highly purified acetyl-CoA carboxylase 2-3-fold at physiological citrate concentrations (0.1-0.5 mM). The effects of the regulator on acetyl-CoA carboxylase activity were dose-dependent, and half-maximal activation occurred in 7-8 min at 30 degrees C. 2. The acetyl-CoA carboxylase regulator was non-dialysable and was inactivated by heating or by exposure to carboxypeptidase. The regulator was enriched from rat liver cytosol by first removing the endogenous acetyl-CoA carboxylase and then using a combination of purification steps, including (NH4)2SO4 precipitation, ion-exchange chromatography and size-exclusion chromatography. The regulator activity appeared to be a protein with a molecular mass of approx. 75 kDa, which could be eluted from mono-Q with approx. 0.35 M KCl as a single peak of activity. 3. Studies of the effects of the regulator on phosphorylation or subunit size of acetyl-CoA carboxylase indicated that the changes in enzyme activity are most unlikely to be explained by dephosphorylation or by proteolytic cleavage. 4. The regulator co-migrates with acetyl-CoA carboxylase through several purification steps, including ion-exchange chromatography and precipitation with (NH4)2SO4; however, the proteins may be separated by Sepharose-avidin chromatography, and the association between the proteins is also disrupted by addition of avidin in solution. Furthermore, the binding of the regulator itself to DEAE-cellulose is altered by the presence of acetyl-CoA carboxylase. Taken together, these observations suggest that the effects of the regulator on acetyl-CoA carboxylase may be explained by direct protein-protein interaction in vitro.
Quayle
K A
KA
Department of Biochemistry, University of British Columbia, Vancouver, Canada.
Denton
R M
RM
Brownsey
R W
RW
D000103:Acetyl-CoA Carboxylase / Q000378:metabolism*; D000818:Animals; D003600:Cytosol / Q000378:metabolism; D004789:Enzyme Activation; D006868:Hydrolysis; D008099:Liver / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D011506:Proteins; D000103:Acetyl-CoA Carboxylase