Below are kinetic data for a Michaelis-Menten enzyme. The total concentration of

Question

Below are kinetic data for a Michaelis-Menten enzyme. The total concentration of enzyme was 1.00 nM (nanomolar, 10-9 M) for each of the five experiments. Perform a Lineweaver-Burk analysis of the data. (Use Microsoft Excel to plot the reciprocal of the initial velocities versus the reciprocal of the substrate concentrations.) From the resulting double reciprocal plot determine the numerical values of Vmax and KM. Finally calculate the turnover number for the enzyme (k2) in units of inverse seconds (s-1). (The turnover number is also referred to as kcat; see page 146 in text.)

Experiment          [S] (µM)              Vo (µM/min)

1                      13                                30

2                      39                                63

3                      79                                87

4                      230                            116

5                      400                            122     

Explanation / Answer

We will plot a graph between:

Vo and Vo /S

So the equation will be:

V0 = Vmax - v0Km / [S]

Equation : y = -46.555x + 137.92

intercept = Vmax = 137.92

Slope = -Km

Km = -slope = 46.555

the enzyme concentration = 10^-9

so turnover number = Vmax / Et = 137.92 / 10^-9 = 137.92 x 10^9

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