Kinetic constants for CoA-derivative substrates of RipA.

CoA transferase activity of RipA was measured with a variety of CoA-derivatives at varying concentrations. Each 100 µL reaction mixture contained CoA-derivative, 50 mM sodium acetate, 100 mM Tris pH 7.0, 1 mM oxaloacetate, 0.5 U citrate synthase and 1 mM DTNB. The reaction was initiated with the addition of 5 µM RipA and was incubated at room temperature for 30 min monitoring the release of free coenzyme A at 412 nm, which detects the formation of the nitrothiobenzoate dianion.