The purification of a recombinant protein is carried out starting with 100 liter
ID: 177336 • Letter: T
Question
The purification of a recombinant protein is carried out starting with 100 liters of a clarified cell lysate, which has a total protein concentration of 0.36 mg/ml and a recombinant prootein concentration of 2.2 U/ml, where U denotes units of biological activity of the recombinant protein. It is known that the completely pure recombinant protein has a specific activity of 40.0 U/mg. Purification is continued until a chromatography step that yields 2.0 liters of a fraction containing the protein, with a total protein concention of 1.11 mg/ml and a recombinant protein concentation of 43.2 U/ml.
For the recombinant protein, calculate the starting and ending specific activity, and the percentage yield and fold purification through the chromatography step.
Explanation / Answer
Total volume of a clarified cell lysate = 100 L or 100000 ml
Total protein concentration ( mg/ml) = 0.36 mg/ml
Total protein (mg) = Protein concentration * total volume
= 0.36 * 100000
= 36000 mg
Total recombinant protein concentration (Activity, units/ml) = 2.2 U/ml
Starting total activity (units) = activity * total volume
= 2.2 * 100000
= 220000 units
Starting specific activity (units/mg) = Total activity / total protein
= 220000 / 36000
= 6.1 units/mg
After purification
Total volume = 2 L or 2000 ml
Total protein concentration ( mg/ml) = 1.11 mg/ml
Total protein (mg) = Protein concentration * total volume
= 1.11 * 2000
= 2220 mg
Total recombinant protein concentration (Activity, units/ml) = 43.2 U/ml
Total activity (units) = activity * total volume
= 43.2 * 2000
= 86400 units
Ending specific activity (units/mg) = Total activity / total protein
= 86400/ 2220
= 39 units/mg
Percentage yield = (Total activity / Starting total activity) * 100
= (86400 /220000 ) * 100
= 40 %
Fold purification = Ending specific activity / starting specific activity
= 39 / 6.1
= 6.4
Related Questions
Navigate
Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.