Explain precipitating protein with ammonium sulfate

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Reference no: EM13646302

An attempt was made to purify a malate dehydrogenase preparation by precipitating protein with ammonium sulfate at 55% saturation. The precipitate was redissolved in buffer, and this solution was found to have a protein concentration of 1.48g/L. A 1:500 dilution was made and aliquots of this were used for the malate

dehydrogenase assay, the initial velocity was 0.11 µmole formed/min and when 15 µL of this diluted enzyme solution was used for the assay, the initial velocity was 0.165 µmole formed/min. The supernatant from the ammonium sulfate precipitation was found to have a protein concentration of 2.05g/L. A 1:1000 dilution was used for the assay. When 10 µL of this diluted enzyme solution was used for the assay, the initial velocity was 0.08 µ mole formed /min and when 15 µL of this diluted enzyme solution was used for the assay, the initial velocity was 0.12 µmole formed/min.

A.Calculate the specific activities of the two fractions.

B. Comment on the usefulness of the ammonium sulphate precipitation as a purification step for this enzyme.

Reference no: EM13646302

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