You discover and purify a new enzyme, and obtain the following data: a) From the

Question

You discover and purify a new enzyme, and obtain the following data: a) From the information in the table, calculate the specific activity (units/mg protein) of the enzyme solution after each purification step. b) Which of the purification procedures used for this enzyme is most effective - gives the greatest increase in purity? c) Which of the purification procedures is least effective? d) Is there any indication from the table that the enzyme is now pure? What else could be done to estimate the purity of the final preparation? e) By how much has the enzyme been purified overall? Assuming that the enzyme is now pure, what percentage of the total protein in the original crude homogenate does the enzyme represent? What assumptions must you make in order to perform this calculation? What are the recoveries of activity after each purification step?

Explanation / Answer

a) i) Specific activity = 4000,000/ 20,000 = 200 units/mg

    ii) Specific activity = 3000,000/5,000 = 600 units/mg

   iii) specific activity = 1000,000/4,000 = 250 units/mg

iv) specific activity = 800,000/200 = 4000 units/mg

v) Specific activity = 750,000/50 = 15000 units/mg

vi) Specific activity = 675,000/45 = 15000 units/mg

b) Affinity chromatopgraphy and Gel-filtration chromatography are most effective as more number of the units get purufied per mg of protien

c) Crude homogenate is the least effective as units/mg of the protein is low.

d) Affinity chromatopgraphy and Gel-filtration chromatography indicate that the enzyme is pure now.

answered the first 4 sub questions of the given question.

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