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Homework Help: Enzyme Activity

  1. Sep 9, 2010 #1
    1. The problem statement, all variables and given/known data

    A student purified cox-2. The crude extract contained 12 g of protein and had 23,000 units of cox-2 activity in 800 mL. The final purification using an affinity column resulted in 3 mL that was 2,300 fold pure. What was the activity of 5 microL of the final purification?


    2. Relevant equations

    See below.

    3. The attempt at a solution

    Under my assumption, enzyme units will be represented by micromol/sec/mL for each purification step. Purification is reflected through specific activity. Therefore,

    Crude extract’s specific activity = micromol/sec/mg protein = (23,000 micromol/sec/mL * 800 mL)/(12,000 mg) = 1533.33 micromol/sec/mg

    Fold of purification = final purification’s specific activity / crude extract’s specific activity


    final purification’s specific activity = Fold of purification * crude extract’s specific activity = 2300*1533.33 micromol/sec/mg = 3.53*10^6 micromol/sec/mg

    This part is where I am confused over as total enzyme activity does not include mg of protein.

    Is this setup permissible?

    12 g / x = 800 mL / 3 mL, where x = final purification’s total mass of protein in g??

    If above is true, then:

    X = .045 g = 45 mg

    3.53*10^6 micromol/sec/mg * 45 mg = 1.587*10^8 micromol/sec

    Activity in 3 mL = (1.587*10^8 micromol/sec)/(3 mL) = 5.29*10^7 micromol/sec/mL

    The total enzyme activity in 5 microL of final purification is: 5.29*10^7 micromol/sec/mL * (5*10^-3 mL) = 2.65*10^5 micromol/sec.

    I don’t know if I have my unit definitions correct, so please tell if my setup is correct. If my approach is wrong, then guide me.

    Thank you.
     
  2. jcsd
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