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Title: Biochemical study of cytidine 3',5' cyclic monophosphate phosphodiesterase activity
Author: Bastani, A.
Awarding Body: University College of Swansea
Current Institution: Swansea University
Date of Award: 1995
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Two enzymes capable of hydrolysing cytidine 3',5'-cyclic monophosphate have previously been reported, one (cCMP-specific PDE) with an absolute specificity for cCMP as substrate, and the other (multifunctional PDE) capable of hydrolysing a number of cyclic nucleotide substrates. This phosphodiesterase preparation was capable of hydrolysing different substrates (cyclic CMP > cyclic AMP > cyclic UMP > cyclic GMP) with the highest specific activity toward cyclic CMP as substrate. It was found to contain both the multifunctional and the cyclic CMP-specific phosphodiesterase, as shown by kinetic data, mass spectrometric analysis, and by utilizing isoelectric focusing and polyacrylamide gel electrophoresis. The PDE activity of the preparation was activated by cytidine and mercaptoethanol, inhibited by aspartate and arginine, but was insensitive to calmodulin. It was active in the absence of metal ions but inhibited by addition of Fe2+ and Ca2+ when cyclic CMP was substrate. Various types of inhibition were observed with different effectors. Cyclic CDP-deoxy cyclic CMP, glutamyl cyclic CMP and 3',5'-cyclic AMP produced competitive inhibition; theophylline produced noncompetitive inhibition; and cytidine 2'-monophosphate 3',5'-cyclic monophosphate, 5-CMP, 2',3'-cyclic CMP, 2',3'-cyclic AMP and 3',5'-cyclic UMP produced mixed type inhibition (either competitive-noncompetitive or uncompetitive-noncompetitive inhibition). Molecular modelling of the substrates and effectors was carried out, and it was deduced that the nitrogen atom bonded to carbon atom 4 (N4) and the oxygen atom bonded to carbon atom 5'(5'-O) with an interatomic distance of 8.92A° was crucial to the binding of ligands; good correlation was obtained between this distance and the potency of the effector. Mass spectrometric analysis suggested that both 5'- and 3'-CMP were products of the enzyme preparation's activity upon 3',5'-cCMP.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available