A capillary electrophoretic assay for acetyl coenzyme A carboxylase

Anal Biochem. 2013 Jun 1;437(1):32-8. doi: 10.1016/j.ab.2013.02.005. Epub 2013 Feb 19.

Abstract

A simple off-column capillary electrophoretic (CE) assay for measuring acetyl coenzyme A carboxylase holoenzyme (holo-ACC) activity and inhibition was developed. The two reactions catalyzed by the holo-ACC components, biotin carboxylase (BC) and carboxyltransferase (CT), were simultaneously monitored in this assay. Acetyl coenzyme A (CoA), malonyl-CoA, adenosine triphosphate (ATP), and adenosine diphosphate (ADP) were separated by capillary electrophoresis, and the depletion of ATP and acetyl-CoA as well as the production of ADP and malonyl-CoA were monitored. Inhibition of holo-ACC by the BC inhibitor, 2-amino-N,N-dibenzyloxazole-5-carboxamide, and the carboxyltransferase inhibitor, andrimid, was confirmed using this assay. A previously reported off-column CE assay for only the CT component of ACC was optimized, and an off-column CE assay for the BC component of ACC also was developed.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Acetyl Coenzyme A / metabolism
  • Acetyl-CoA Carboxylase / antagonists & inhibitors
  • Acetyl-CoA Carboxylase / metabolism*
  • Adenosine Triphosphate / metabolism
  • Biocatalysis
  • Carbon-Nitrogen Ligases / metabolism
  • Carboxyl and Carbamoyl Transferases / metabolism
  • Electrophoresis, Capillary / methods*
  • Enzyme Assays / methods*
  • Enzyme Inhibitors / pharmacology
  • Escherichia coli / enzymology
  • Holoenzymes / metabolism

Substances

  • Enzyme Inhibitors
  • Holoenzymes
  • Acetyl Coenzyme A
  • Adenosine Triphosphate
  • Carboxyl and Carbamoyl Transferases
  • Carbon-Nitrogen Ligases
  • biotin carboxylase
  • Acetyl-CoA Carboxylase