Regulation of enzymes by fatty acyl coenzyme A. Site-specific binding of fatty acyl coenzyme A by citrate synthase–a spin-labeling study.

Abstract

4.1.3.7) at about the same concentration as did palmitoyl-CoA. Competitive binding studies show acetyl-CoA and ATP to displace 6- and 16-doxylstearoyl-CoA from citrate synthase but little while oxalacetate and NADPH compete very effectively. These studies may suggest binding by fatty acyl-CoA at sites apart from the acetyl-CoA binding site. The differential displacement by ATP and NADPH argues against binding of the fatty acyl-CoA at a general nucleotide binding site. The spectrum of 6-doxylstearoyl-CoA in the presence of high concentrations of citrate synthase was for an immobilized spin label; under the same conditions the spectrum for 16-doxylstearoyl-CoA was mobile. These spectral differences were interpreted in terms of a specific binding site for fatty acyl-CoA in which the portion of the fatty acyl chain proximal to the thioester was immobilized while the methyl end was allowed greater motional freedom. Phospholipid vesicles included the spin label of 16-doxylstearoyl-CoA but not that of 6doxylstearoyl-CoA in the presence of excess citrate synthase in support of the above interpretation. A model for a specific binding site for fatty acyl-CoA on citrate synthase with provision for interaction with a biological membrane is presented. Site-specific binding of fatty acyl-CoA by citrate synthase provides strong support for a physiological role for fatty acyl-CoA in the regulation of citrate synthase and by analogy for other related enzymes of lipid metabolism.