Enzyme Kinetics - Effect of Enzyme Concentration and Substrate

Instructions:

  1. Select an enzyme concentration that will give reading within range, when substrate is saturating.
  2. Keeping the enzyme concentration fixed, vary [S] and measure the absorption, obtain 5-6 values.
  3. Use an extinction coefficient of 18,300 M-1cm-1 to convert the absorption to product.
  4. Plot the amount of product produced/sec versus [S] and determine KM, VMax, and kCAT.

Note that your instrument cannot read absorbance values greater than 2.

1. Enter a value for E_total in this box:
ETOT = nM
2. Enter a value for [S] in this box:
[S] = mM
3. For the above values of ETot and [S], Calculate Absorbance/time
A/sec =
    (Each calculated value has a small "experimental error" added to it.)

10.9.2020