Question

Given the information that follows, complete the purification table. A 50 mL crude extract of skeletal muscle contains 32 mg of protein per mL. Ten μL of the extract catalyzes a reaction at a rate of 0.14 μmol product formed per minute. The extract was fractionated by ammonium sulfate precepitation, and the fraction precipitating between 20% and 40% saturation was redissolved in 10 mL. This solution contains 50 mg/mL protein. Ten μL of the purified fraction catalyzes the reaction at a rate of 0.65 μmol/min.

Need to find the Crude and Purified values of the following:

Total Protein (mg)	Specific Activity (µmol·min-1·µL-1·mg-1)	Total Activity (µmol·min-1)	% Yield	Fold Purification
Crude
Purified

Answer 1

1st case-

Crude extract-

50 mL crude extract contains 32 mg of protein per mL. Ten μL of the extract catalyzes a reaction at a rate of 0.14 μmol product formed per minute.

Rate = 0.14 umol/min

Concentration of crude extract= 32mg/ml

Thus amount of extract in 10ul = 32*10/1000 mg= 0.32 mg

Thus the Specific activity of the crude= rate/amount of extract used= 0.14/0.32 = 0.437 umol/min/mg

2nd case-

Purified protein-

Rate= 0.65 μmol/min

Concentration= 50 mg/mL

Amount of protein in 10ul = 50*10/1000 mg= 0.5mg

Thus the specific activity of the purified protein= Rate/amount of protein= 0.65/0.5 = 1.3 umol/min/mg