Unique structural features and differential phosphorylation of the 280-kDa component (isozyme) of rat liver acetyl-CoA carboxylase

J Biol Chem. 1994 May 20;269(20):14438-45.

Abstract

Rat liver acetyl-CoA carboxylase (ACC, EC 6.4.1.2) exhibits major and minor subunits (M(r) of 265,000 and 280,000 respectively), the structure and function of which are compared in this study. The two subunits copurified and each contained biotin as demonstrated by avidin reactivity and direct determination of biocytin. In agreement with previous studies, the ACC subunits could be distinguished with specific monoclonal antibodies and differential tissue expression. We now report extensive differences in primary structure revealed by peptide mapping, mass spectrometric analysis of peptides following reverse phase high performance liquid chromatography, and microsequencing of selected peptides. Four peptides derived from the 265-kDa subunit were sequenced and matched sequences within the predicted structure of rat 265-kDa ACC. Although one identical peptide sequence was detected within both subunits (residues 2009-2024 of the 265-kDa subunit), 12 peptides derived from the 280-kDa subunit exhibited entirely novel sequences or matched partially (average 70% identity) with sequences within the 265-kDa subunit. The 280-kDa subunit may also exhibit distinct functional properties, since the initial rate of phosphorylation was at least 10-fold greater than that of the 265-kDa subunit in the presence of cAMP-dependent protein kinase. Two-dimensional mapping demonstrated that the tryptic phosphopeptides released from the two ACC subunits are distinct. These structural studies suggest that the 265- and 280-kDa components (isozymes) of ACC are so distinct they may be encoded by separate genes, while the differential phosphorylation observed in vitro suggests a key role for the 280-kDa subunit in regulating enzyme activity within intact cells.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetyl-CoA Carboxylase / chemistry*
  • Acetyl-CoA Carboxylase / isolation & purification
  • Acetyl-CoA Carboxylase / metabolism*
  • Amino Acid Sequence
  • Animals
  • Biotin / analysis
  • Chromatography, High Pressure Liquid
  • Electrophoresis, Polyacrylamide Gel
  • Isoenzymes / chemistry*
  • Isoenzymes / isolation & purification
  • Isoenzymes / metabolism*
  • Liver / enzymology*
  • Lysine / analogs & derivatives
  • Lysine / analysis
  • Macromolecular Substances
  • Mass Spectrometry
  • Molecular Sequence Data
  • Molecular Weight
  • Peptide Fragments / chemistry
  • Peptide Fragments / isolation & purification
  • Peptide Mapping
  • Phosphopeptides / chemistry
  • Phosphopeptides / isolation & purification
  • Phosphorylation
  • Rats
  • Rats, Wistar
  • Sequence Homology, Amino Acid

Substances

  • Isoenzymes
  • Macromolecular Substances
  • Peptide Fragments
  • Phosphopeptides
  • Biotin
  • Acetyl-CoA Carboxylase
  • biocytin
  • Lysine