nd W and in the process produces a faint yellow glow, so assaying Your enzyme ox

Question

nd W and in the process produces a faint yellow glow, so assaying Your enzyme oxidizes compou activity of your enzyme is easy. You are ready to purify your enzyme. You begin with a crude lysate of bacteria, and after centrifugation, you find activity only in the supernatant. You first 8- precipitate proteins out of the crude lysate supernatant at 15% ammonium sulfate, but there was no activity in the 15% ammonium sulfate pellet. You increase the amount of ammonium, sulfate in the supernatant from 15% to 25% but there was no activity in the 25% ammonium sulfate pellet. You increase the amount of ammonium sulfate in the supernatant from 25% to 45%, and now all of the activity is in the pellet. In the lab, you precipitated -galactosidase out of the crude lysate supernatant at 45% ammonium sulfate. Compare the precipitation above for enzyme X and the one in the lab for -gal, was one a better purification than the other or were they the same? Explain your answer

Explanation / Answer

The purification of the enzyme X is better compared to the purification of b galactosidase because when isolating b galactosidase, 45% of ammonium sulfate is directly used to isolate the protein b galactosidase. So, in this case all the proteins that are precipitated between 0-45 percent are isolated at the same time in the pellet.

But, in case of protein X, the concentration is gradually increased and the pellet in which the Protein X is observed is having proteins that are precipitated in the range of 25-45 % ammonium sulfate. So, the pellet is containing higher proportion of Protein X compared to b galactosidase in same pellet amount. i.e. Protein X is more purified than b Galactosidase.

Thank you.

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