Quantitation by fast‐atom bombardment/mass‐analysed ion kinetic energy spectrometry: Kinetic analysis of cyclic nucleotide phosphodiesterase activity
- 1 June 1989
- journal article
- research article
- Published by Wiley in Rapid Communications in Mass Spectrometry
- Vol. 3 (6) , 178-182
- https://doi.org/10.1002/rcm.1290030604
Abstract
Quantitation of cyclic nucleotide phosphodiesterase activity by means of fast‐atom bombardment (FAB) mass spectrometry with mass‐analysed ion kinetic energy (MIKE) spectrum scanning is described. Characteristic peaks of the substrate, cyclic AMP, and product, AMP, were identified in positive‐ion FAB mass spectra and MIKE scans of the protonated molecules. By spiking enzyme incubates with known quantities of cyclic AMP and AMP and measuring peak heights in the MIKE spectra of both spiked and unspiked samples, the concentrations of cyclic AMP and AMP in solution at the end of a series of enzyme incubations have been estimated. From the data obtained the Km and Vmax of the enzymes were calculated as 181 μM and 28.6 nmol/min respectively, showing excellent agreement with values of the Michaelis constant, Km=205 μM and the maximum velocity Vmax=33.2 nmol/min obtained by radioactive assay.Keywords
This publication has 12 references indexed in Scilit:
- Cytidylate cyclase activity: Identification of cytidine 3′,5′‐cyclic monophosphate and four novel cytidine cyclic phosphates as biosynthetic products from cytidine triphosphateRapid Communications in Mass Spectrometry, 1988
- Enzymes and mass spectrometry: A dynamic combinationMass Spectrometry Reviews, 1987
- Determination of Km and Vmax for tryptic peptide hydrolysis using fast atom bombardment mass spectrometryAnalytical Chemistry, 1986
- The differentiation of isomeric biological compounds using collision-induced dissociation of ions generated by fast atom bombardmentJournal of Mass Spectrometry, 1985
- Extraction, purification and identification of cytidine 3′,5′-cyclic monophosphate from rat tissuesBiochemical Journal, 1984
- The identification of cyclic nucleotides from living systems using collision-induced dissociation of ions generated by fast atom bombardment mass spectrometryJournal of Mass Spectrometry, 1984
- Occurrence of guanosine 3′,5′-cyclic monophosphate (Cyclic GMP) and associated enzyme systems in Phaseolus vulgarisPhytochemistry, 1984
- Identification of endogenous cytidine 3′:5′ -cyclic monophosphateBiochemical Society Transactions, 1983
- [30] Assay of cyclic nucleotide phosphodiesterases with radioactive substratesPublished by Elsevier ,1974
- The Determination of Enzyme Dissociation ConstantsJournal of the American Chemical Society, 1934