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Biology LibreTexts

15: Ribozymes, Enzyme Kinetics

Reading& Problems: LNC p. 200-206; p. 238 prob. 8, 11, 13; Segel, p.319, prob. 1,2,3

I. Enzyme Kinetics - measuring/calculating the Velocity of an enzyme catalyzed reaction:

Rate of production of product is the velocity V = d[P]/dt

A. Assumptions

 

  • Initial rate - we are in the initial phase of the reaction when the reaction has proceeded to a small enough extent that the [S] does not change significantly.
  • Steady state - the [ES] rapidly reaches a steady state that does not change over the time being analyzed.

 

B. From these two assumptions we can derive the Michaelis-Menten equation for enzyme kinetics.

 

C. Define Vmax as the maximal rate at a given enzyme concentration, Vmax = kcat[Et]

 

D. Plotting enzyme data (here is a link to download external link: the enzyme plots used in class)

 

  • [P] vs. time
  • V versus [S]

 

II. Enzyme kinetics, continued.

 

A. Lineweaver-Burk double reciprocal plot: 1/V vs 1/[S] and determination of Km (X-intercept is -1/Km) and Vmax (Y-intercept is 1/Vmax), slope is Km/Vmax

 

B. The meaning of the constants

 

  • Km - the [S] that leads to half maximal velocity, a measure of the affinity of the enzyme for the substrate. Range 0.4 µM - 10 mM. Lower Km means higher affinity.
  • Vmax - not really a constant, =kcat[Et] and [Et] can vary. It is only a constant at a constant enzyme concentration.
  • Kcat - how many reactions one enzyme can perform per unit time. In inverse time units. In sec-1 (per second) called "turnover number". Range from less than 0.5 up to 6 x 105. Higher Kcat means faster reaction.
  • Best measure of enzyme efficiency is Kcat/Km. Bigger number is more efficient enzyme.

Some take home information:
Enz Kinet Eqn.jpg
Lineweaver-Burk double reciprocal plot: 1/V vs 1/[S] can be used to determine Km (X-intercept is -1/Km) and Vmax (Y-intercept is 1/Vmax)

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