Abstract
Methods are described for the assay of ATP citrate ( pro-3S)-lyase (EC 4.1.3.8) and acetyl-CoA synthetase (EC 6.2.1.1) activity in crude tissue extracts. Arylamine acetyltransferase (EC 2.3.1.5) and p-nitroaniline serve as indicator system in these tests. The methods have several advantages with respect to other optical enzyme assays. They are more sensitive than NADH- or hydroxylamine-dependent methods due to the higher molar extinction coefficient of p-nitroaniline, which was redetermined under assay conditions. The assay is independent of nonspecific “NADH oxidase” activity. Continuous readings of the optical density can be performed. Specific activities of ATP citrate-lyase and acetyl-CoA synthetase have been measured in livers of starved and refed rats. Carbohydraterich refeeding leads to an increase in both enzyme activities. A modification of the biuret method for the use in lipid-rich tissue extracts is described.
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