Biochemistry I Fall Term

Determination of KI using a Secondary Plot

  1. Double Reciprocal Plot: 1/vo vs. 1/[A].

    The above graph is the same as that on the Enzyme Inhibition: Answer Sheet page. The bottom (blue) line corresponds to the data without added I. The intercept and slope yielded, Vmax = 0.5 mM/min and KM = 2 mM.
    The top two lines (green and red) correspond to the lines with 2 mM and 5 mM added [I], respectively.
  2. Secondary Plot: Slopes vs. [I].

    The slopes from the above double reciprocal plot are plotted in a secondary plot (or "replot") vs. the [I]'s used.
    The intercept of this graph is, KM/Vmax.
    The slope of this graph is, KM/(VmaxKI).
    Thus, KI = Int/Slope = 3 mM.

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