Question

What is the next step after organic solvent fractionation with dry ice – ethanol – water mixture?

(a) Refrigerated centrifugation at 3000 g for 15 mins

(b) Drying in freezer

(c) Supernatant fractionation with ethanol

(d) Precipitated protein dissolved in 0.01 to 0.05 M cold buffer

I got this question during a job interview.

My query is from Enzymes Purification in section Enzymes Purification of Enzyme Technology

Answer 1

The correct answer is (a) Refrigerated centrifugation at 3000 g for 15 mins

To explain I would say: In organic solvent fractionation, the desired protein to be purified is kept a bath containing dry ice – ethanol -water mixture. Constantly ethanol is added which increases temperature of the solution. While adding the ethanol the protein solution is stirred continuously and also after all the ethanol is added for 15 mins. Then the solution is centrifuged in refrigerated centrifugation at 3000 g for 15 mins. The resultant supernatant is fractionated with ethanol, followed by drying in freezer keeping the tubes in inverted position. The last step is to dissolve the precipitated protein with 0.01 to 0.05 M cold buffer. Hence the first step after organic solvent fractionation is refrigerated centrifugation at 3000 g for 15 mins.