title: Catalysis by entropic effects: the action of cytidine deaminase on 5,6-dihydrocytidine.
abstract: In neutral solution, 5,6-dihydrocytidine undergoes spontaneous deamination (k25 approximately 3.2 x 10(-5) s(-1)) much more rapidly than does cytidine (k25 approximately 3.0 x 10(-10) s(-1)), with a more favorable enthalpy of activation (DeltaDeltaH# = -8.7 kcal/mol) compensated by a less favorable entropy of activation (TDeltaDeltaS# = -1.8 kcal/mol at 25 degrees C). E. coli cytidine deaminase enhances the rate of deamination of 5,6-dihydrocytidine (kcat/k(non) = 4.4 x 10(5)) by enhancing the entropy of activation (DeltaDeltaH# = 0 kcal/mol; TDeltaDeltaS# = +7.6 kcal/mol, at 25 degrees C). Binding of the competitive inhibitor 3,4,5,6-tetrahydrouridine (THU), a stable analogue of 5,6-dihydrocytidine in the transition state for its deamination, is accompanied by a release of enthalpy (DeltaH = -7.1 kcal/mol, TDeltaDeltaS = +2.2 kcal/mol) that approaches the estimated enthalpy of binding of the actual substrate in the transition state for deamination of 5,6-dihydrocytidine (DeltaH = -8.1 kcal/mol, TDeltaDeltaS = +6.0 kcal/mol). Thus, the shortcomings of THU in capturing all of the binding affinity expected of an ideal transition-state analogue reflect a less favorable entropy of association. That difference may arise from the analogue's inability to displace a water molecule from the "leaving group site" at which ammonia is generated in the normal reaction. The effect on binding of removing the 4-OH group from the transition-state analogue THU, to form 3,4,5,6-tetrahydrozebularine (THZ) (DeltaDeltaH = -2.1 kcal/mol, TDeltaDeltaS = -4.4 kcal/mol), is mainly entropic, consistent with the inability of THZ to displace water from the "attacking group site". These results are consistent with earlier indications [Snider, M. J., and Wolfenden, R. (2001) Biochemistry 40, 11364] that site-bound water plays a prominent role in substrate activation and inhibitor binding by cytidine deaminase.
authors.affiliation: Department of Biochemistry & Biophysics and Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina 27599-7260, USA.
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Snider
authors.forename: Danijela
authors.initials: D
authors.lastname: Lazarevic
authors.forename: Richard
authors.initials: R
authors.lastname: Wolfenden
mesh_terms: D000588:Amines / Q000378:metabolism; D002384:Catalysis; D003562:Cytidine / Q000031:analogs & derivatives / Q000378:metabolism*; D003564:Cytidine Deaminase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019277:Entropy; D004791:Enzyme Inhibitors / Q000378:metabolism
chemical_list: C451739:5,6-dihydrocytidine; D000588:Amines; D004791:Enzyme Inhibitors; D003562:Cytidine; D003564:Cytidine Deaminase