Wk6RG
Sunday, February 10, 2019 8:31 PM
•
RGWk6
• Enzyme kinetics: an approach that involves determining the rate of the (catalyzed?)
reaction and how it changes in response to experimental parameter changes.
• Once the substrate reaches a high enough conc, the max velocity will be reached b/c the
product conc has reached a sufficient amount. Enzyme is "saturated" w/ the substrate ->
further increase in [S] will have no effect on rate. (mostly in the form of ES complex) Vmax
= [ES] =Etotal
• The overall rate must be proportional to the conc of the ES complex b/c the step where the
ES complex breaks down (complex is the reactant) is slower, thus limiting/defining the
reaction rate.
• Steady state: the state in which [ES] remains constant over time. When intermediate ES
remains steady, P is generated at the same time S is consumed.
• Michaelis-Menten Eqn: V0 = Vmax [S] / (Km + [S])
• [P] is negligible early in the rxn, reverse rxn P->S can be ignored.
• Steady-state assumption: the rate of ES formation is equal to ES breakdown.
○ W/o this assumption, we cannot determine the Michaelis constant (Km).
• Initial velocity is dependent on [S].
○ will reach max velocity at high [S].
○ If V0=1/2Vmax, then Km=[S]. Holds for all enzymes that follow Michaelis-Menten
kinetics.
• Also know half of the enzymes are bound to the substrate. (Vmax = [E])
• Parameters Vmax and Km can be found experimentally for any given enzyme. Provide
little info on the steps of the mechanism. Many enzymes that follow Michaelis-Menten
kinetics have a different mechanism than:
E+S ES E+P
○ Vmax = k2[Et], k2 is rate-limiting step
• Double-Reciprocal plot: used to determine a more accurate Vmax.
1/V0 = Km/Vmax[S] + 1/Vmax
• Kcat: the limiting-rate constant of any enzyme-catalyzed rxn at saturation. (kcat = k2)
○ Turnover number: the # of substrate molecules converted to product in a given unit
of time on a single enzyme molecule when the enzyme is saturated w/ substrate.
• Specificity constant: rate constant for the conversion of E + S to E + P.
Km: Michaelis-Menten
constant which shows the
concentration of the substrate when
the reaction velocity is equal to one
half of the maximal velocity.
Sunday, February 10, 2019 8:31 PM
•
RGWk6
• Enzyme kinetics: an approach that involves determining the rate of the (catalyzed?)
reaction and how it changes in response to experimental parameter changes.
• Once the substrate reaches a high enough conc, the max velocity will be reached b/c the
product conc has reached a sufficient amount. Enzyme is "saturated" w/ the substrate ->
further increase in [S] will have no effect on rate. (mostly in the form of ES complex) Vmax
= [ES] =Etotal
• The overall rate must be proportional to the conc of the ES complex b/c the step where the
ES complex breaks down (complex is the reactant) is slower, thus limiting/defining the
reaction rate.
• Steady state: the state in which [ES] remains constant over time. When intermediate ES
remains steady, P is generated at the same time S is consumed.
• Michaelis-Menten Eqn: V0 = Vmax [S] / (Km + [S])
• [P] is negligible early in the rxn, reverse rxn P->S can be ignored.
• Steady-state assumption: the rate of ES formation is equal to ES breakdown.
○ W/o this assumption, we cannot determine the Michaelis constant (Km).
• Initial velocity is dependent on [S].
○ will reach max velocity at high [S].
○ If V0=1/2Vmax, then Km=[S]. Holds for all enzymes that follow Michaelis-Menten
kinetics.
• Also know half of the enzymes are bound to the substrate. (Vmax = [E])
• Parameters Vmax and Km can be found experimentally for any given enzyme. Provide
little info on the steps of the mechanism. Many enzymes that follow Michaelis-Menten
kinetics have a different mechanism than:
E+S ES E+P
○ Vmax = k2[Et], k2 is rate-limiting step
• Double-Reciprocal plot: used to determine a more accurate Vmax.
1/V0 = Km/Vmax[S] + 1/Vmax
• Kcat: the limiting-rate constant of any enzyme-catalyzed rxn at saturation. (kcat = k2)
○ Turnover number: the # of substrate molecules converted to product in a given unit
of time on a single enzyme molecule when the enzyme is saturated w/ substrate.
• Specificity constant: rate constant for the conversion of E + S to E + P.
Km: Michaelis-Menten
constant which shows the
concentration of the substrate when
the reaction velocity is equal to one
half of the maximal velocity.
