title: The complete pathway for ERK2-catalyzed reaction. Evidence for an iso random Bi Bi mechanism.
abstract: In the present study, the enzymatic mechanism of ERK2 is re-examined by a combination of steady-state kinetic studies in the absence and presence of viscosogenic agents. Kinetic studies carried out in various concentrations of sucrose revealed that both k(cat) and k(cat)/K(m) for either ATP or EtsDelta138 were highly sensitive to solvent viscosity, suggesting that the rapid equilibrium assumption is not valid for the phosphorylation of protein substrate by ERK2. Furthermore, the kinetic analysis with the minimal random Bi Bi reaction mechanism is shown to be inconsistent with the principle of the detailed balance. This inconsistent calculation strongly suggests that there is isomerization of the enzyme-substrate ternary complex. The viscosity-dependent steady-state kinetic data are combined to establish a kinetic mechanism for the ERK2-catalyzed reaction that predicts initial reaction velocities under varying concentrations of ATP and substrate. These results complement previous structure-function studies of mitogen-activated protein kinases and provide important insight for mechanistic interpretation of the kinase functions.
authors.affiliation: Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China. zxwang@sun5.ibp.ac.cn
authors.forename: Zhi-Xin
authors.initials: ZX
authors.lastname: Wang
authors.forename: Jia-Wei
authors.initials: JW
authors.lastname: Wu
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry*; D002384:Catalysis; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D008956:Models, Chemical; D008962:Models, Theoretical; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D012997:Solvents / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D014783:Viscosity
chemical_list: D012997:Solvents; D000255:Adenosine Triphosphate; D019950:Mitogen-Activated Protein Kinase 1