Kinetic analysis of cyclic CMP-specific and multifunctional phosphodiesterases by quantitative positive-ion fast-atom bombardment mass spectrometry

Two enzymes, cyclic CMP-specific phosphodiesterase and multifunctional phos phodiesterase, are responsible for the hydrolysis of cytidine: 3',5'-cyclic monophosphate in living cells. Quantitation of both enzymes has been carri ed out by positive-ion fast-atom bombardment mass spectrometric analysis of the enzyme incubates after termination of the reaction. The kinetic data o btained are in close agreement with parallel data obtained by the conventio nal radiometric assay. The extra facility of the mass spectrometry based as say to monitor several incubation components simultaneously has been exploi ted to study the concurrent hydrolysis of alternate cyclic nucleotide subst rates and provides kinetic parameters of significance in interpreting subst rate-enzyme interactions. This is extended by the use of collisionally-indu ced dissociation of the protonated molecules of the liberated products to i dentify the mononucleotide Isomers resulting from the cyclic nucleotide hyd rolysis. Copyright (C) 1999 John Wiley & Sons, Ltd.