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Kinetic analysis of cyclic CMP‐specific and multifunctional phosphodiesterases by quantitative positive‐ion fast‐atom bombardment mass spectrometry
Author(s) -
Newton Russell P.,
Bayliss Mark A.,
Khan Jalal A.,
Bastani Abdolhossein,
Wilkins Adam C. R.,
Games David E.,
Walton Terence J.,
Brenton A. Gareth,
Harris Frank M.
Publication year - 1999
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/(sici)1097-0231(19990415)13:7<574::aid-rcm526>3.0.co;2-r
Subject(s) - chemistry , fast atom bombardment , mass spectrometry , protonation , hydrolysis , dissociation (chemistry) , kinetic energy , substrate (aquarium) , enzyme , ion , molecule , phosphodiesterase , collision induced dissociation , nucleotide , chromatography , organic chemistry , tandem mass spectrometry , biochemistry , physics , oceanography , quantum mechanics , gene , geology
Two enzymes, cyclic CMP‐specific phosphodiesterase and multifunctional phosphodiesterase, are responsible for the hydrolysis of cytidine 3′,5′‐cyclic monophosphate in living cells. Quantitation of both enzymes has been carried out by positive‐ion fast‐atom bombardment mass spectrometric analysis of the enzyme incubates after termination of the reaction. The kinetic data obtained are in close agreement with parallel data obtained by the conventional radiometric assay. The extra facility of the mass spectrometry based assay to monitor several incubation components simultaneously has been exploited to study the concurrent hydrolysis of alternate cyclic nucleotide substrates and provides kinetic parameters of significance in interpreting substrate–enzyme interactions. This is extended by the use of collisionally‐induced dissociation of the protonated molecules of the liberated products to identify the mononucleotide isomers resulting from the cyclic nucleotide hydrolysis. Copyright © 1999 John Wiley & Sons, Ltd.