Question

(a) After spending a few weeks isolating the enzyme, you have enough pure enzyme to do some kinetic experiments, using a chromogenic substrate. The product produced by the enzyme from this substrate has an extinction coefficient of 20,500cm−1M−1 at 486nm, whereas the substrate does not absorb light significantly at this wavelength. For these experiments, you have used an enzyme concentration of 1.75µg/mL and monitored the change of absorbance at 486nm, using cuvettes with a path length of 1cm. The initial results are summarized in the table below:
[S] (µM) ∆ A/min
0 0.0
50 0.020
100 0.028
250 0.036
500 0.040
750 0.042
1000 0.043
From the data above, estimate the value of Km, in µM units, and Vmax, in µM/min. You should not have to do any graphing, curve-fitting or extensive calculations to estimate these parameters!

Answer 1