Malonyl-CoA: acetyl-CoA cycling. A new micromethod for determination of acyl-CoAs with malonate decarboxylase.

A new micromethod for determination of acetyl-CoA and malonyl-CoA using malonate decarboxylase is described. This enzyme catalyzes decarboxylation of malonate in a cyclic manner and produces acetate in proportion to the amount of a given acyl-CoA, such as acetyl-CoA, malonyl-CoA or propionyl-CoA. The acetate generated is converted to acetylphosphate by acetate kinase (EC 2.7.2.1) added at the same time and is determined spectrophotometrically as acetohydroxamate. The sensitivity of this method is high enough to detect 10(-12) mol of acetyl-CoA or malonyl-CoA. The simplicity of the method allows more than 30 samples to be analyzed at the same time without any prior extraction step. Although this method does not distinguish between acetyl-CoA and malonyl-CoA, malonyl-CoA alone can be measured by elimination of acetyl-CoA with citrate synthetase (EC 4.1.3.7).