A general coupled spectrophotometric assay for decarboxylases.
A rapid, continuous spectrophotometric method has been developed for the assay of decarboxylases. The assay uses a coupled enzyme system in which liberated CO2 is reacted with phosphoenolpyruvate and phosphoenolpyruvate carboxylase to form oxaloacetate, which in turn is reduced by malate dehydrogenase to L-malate concomitantly with the oxidation of NADH to NAD. The resultant decrease in absorbance at 340 nm accurately reflects the activity of the decarboxylase. The method is capable of detecting the liberation of as little as 1 nmol of CO2/min and was tested in assays of lysine decarboxylase, orotidine-5'-phosphate decarboxylase, and 4'-phosphopantothenoyl-L-cysteine decarboxylase.
D002245:Carbon Dioxide / Q000302:isolation & purification; D002262:Carboxy-Lyases / Q000032:analysis*; D055598:Chemical Phenomena; D002621:Chemistry; D003653:Decarboxylation; D008293:Malates / Q000302:isolation & purification; D009097:Multienzyme Complexes; D009243:NAD; D010084:Oxidation-Reduction; D010453:Peptide Synthases; D013053:Spectrophotometry / Q000379:methods
D008293:Malates; D009097:Multienzyme Complexes; D009243:NAD; D002245:Carbon Dioxide; C030298:malic acid; D002262:Carboxy-Lyases; C021520:lysine decarboxylase; C022373:Phosphopantothenoyl-cysteine decarboxylase; D010453:Peptide Synthases