Protein-serine kinase from rat epididymal adipose tissue which phosphorylates and activates acetyl-CoA carboxylase. Possible role in insulin action.
1. Most of the cyclic-nucleotide-independent acetyl-CoA carboxylase kinase activity in an extract of rat epididymal adipose tissue was evaluated from a Mono Q column by 0.175 M-NaCl at pH 7.4. The activity of the kinase in this fraction (fraction 1) was increased after exposure of intact tissue to insulin. 2. Incubation of purified adipose-tissue acetyl-CoA carboxylase with [gamma-32P]ATP and samples of fraction 1 led to the incorporation of up to 0.4 mol of 32P/mol of enzyme subunit. Most of the phosphorylation was on serine residues within a single tryptic peptide. This peptide, on the basis of two-dimensional t.l.c. analysis, h.p.l.c. and Superose 12 chromatography, appeared to be the same as the acetyl-CoA carboxylase peptide ('I'-peptide) which exhibits increased phosphorylation in insulin-treated tissue. 3. Phosphorylation of purified acetyl-CoA carboxylase by the kinase in fraction 1 was found to be associated with a parallel 4-fold increase in activity. However, increases in both phosphorylation and activity were much diminished if fraction 1 was treated by Centricon centrifugation to remove low-Mr components. Among these components was a potent inhibitor of acetyl-CoA carboxylase activity which appeared to be necessary for the kinase in fraction 1 to be fully active. 4. The inhibitor remains to be identified, but inhibition requires MgATP, although the inhibitor itself does not cause any phosphorylation of the carboxylase. No effects of insulin were observed on the activity of the inhibitor. 5. It is concluded that the kinase probably plays an important role in the mechanism whereby insulin brings about the well-established increases in phosphorylation and activation of acetyl-CoA carboxylase in adipose tissue.
Borthwick
A C
AC
Department of Biochemistry, School of Medical Sciences, University of Bristol, U.K.
Edgell
N J
NJ
Denton
R M
RM
D000103:Acetyl-CoA Carboxylase / Q000378:metabolism*; D000273:Adipose Tissue / Q000201:enzymology*; D000595:Amino Acid Sequence; D000818:Animals; D002845:Chromatography; D004789:Enzyme Activation; D004822:Epididymis / Q000201:enzymology; D007328:Insulin / Q000502:physiology; D008297:Male; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism*; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D051381:Rats; D011919:Rats, Inbred Strains
D007328:Insulin; D010455:Peptides; D010768:Phosphoserine; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D000103:Acetyl-CoA Carboxylase