Surface-mediated enzymatic reactions: simulations of tissue factor activation of factor X on a lipid surface.
Blood coagulation proceeds via reactions in which zymogen coagulation factors are activated to proteases. An essential step is the activation of factor X by a complex of tissue factor and factor VIIa. This complex usually is studied using phospholipid vesicles into which tissue factor is inserted. Because factor X exists free in solution and bound to the lipid-surface, it is difficult to establish experimentally the kinetic contribution of surfaces. We therefore developed a stochastic model to simulate such reactions and generate initial velocity data from which Michaelis-Menten parameters are estimated. Simulated Km values decrease slightly when substrate binding to lipid is increased and by a factor of four when the rates of surface diffusion are increased to that of fluid phase-diffusion. Simulations with various size planar surfaces established an enzyme capture radius of 32-64 nm. Simulations with different modes of enzyme-substrate complex assembly show that if the true substrate is lipid-bound, under certain conditions, the true Kcat is not measured; rather, the product "leaving rate" from the complex is the rate-limiting step that is measured as substrate is taken to infinity. This model is applicable to any surface-bound enzyme reaction.
Department of Mathematics and Statistics, University of Guelph, Ontario, Canada.
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Gentry
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Ye
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Nemerson
D000818:Animals; D001665:Binding Sites; D055592:Biophysical Phenomena; D001703:Biophysics; D001777:Blood Coagulation / Q000502:physiology; D004058:Diffusion; D004798:Enzymes / Q000378:metabolism; D015942:Factor VIIa / Q000378:metabolism; D005170:Factor X / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D008563:Membrane Lipids / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D013379:Substrate Specificity; D013925:Thromboplastin / Q000378:metabolism*
D004798:Enzymes; D008051:Lipid Bilayers; D008563:Membrane Lipids; D005170:Factor X; D013925:Thromboplastin; D015942:Factor VIIa