title: Human deoxycytidine kinase: kinetic mechanism and end product regulation.
abstract: The kinetic properties of the monomeric deoxycytidine kinase (EC 2.7.1.74) from leukemic human T-lymphoblasts have been investigated. The results of steady-state initial-rate kinetic analysis and product inhibition studies at pH 7.5 and 37 degrees C indicate that substrate binding follows an ordered sequential pathway, with the magnesium salt of ATP being the first substrate to bind and dCMP the last product to dissociate. At subsaturating substrate concentrations, dCMP produced competitive inhibition against ATP, while against varied deoxycytidine concentrations dCMP exhibited mixed-type inhibition. ADP produced noncompetitive inhibition against either substrate. The limiting Km values for deoxycytidine and MgATP were 0.94 and 30 microM, respectively. The end product inhibitor dCTP exhibited competitive inhibition against varied ATP concentration, with a dissociation constant estimated to be 0.7 microM when extrapolated to zero ATP concentration. dCTP was purely noncompetitive against varied deoxycytidine concentration. On the basis of these kinetic results, and on the strong and specific inhibition by dCTP, it is proposed that this end product functions as a multisubstrate analogue, with its triphosphate group binding to the phosphate donor site of the enzyme and its deoxycytidine moiety overlapping and binding to the deoxynucleoside site in a highly specific manner.
authors.affiliation: Department of Biochemistry, Ohio State University, Columbus 43210.
authors.forename: M Y
authors.initials: MY
authors.lastname: Kim
authors.forename: D H
authors.initials: DH
authors.lastname: Ives
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D003841:Deoxycytidine / Q000494:pharmacology; D003842:Deoxycytidine Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D010770:Phosphotransferases / Q000378:metabolism*; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured
chemical_list: D003841:Deoxycytidine; D000244:Adenosine Diphosphate; D010770:Phosphotransferases; D003842:Deoxycytidine Kinase