Biochemistry

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Leupeptin and Enzyme Purification

BIOCHM-RHOKSY

A student was assigned the task of purifying an enzyme from liver tissue. They prepared their extraction solution but forgot to add leupeptin (see structure below). Their normally stable enzyme now displayed no activity after the first extraction step.

Edgar181. "Leupeptin." Wikimedia Commons. N.p., 20 June 2011. Web. 07 Aug. 2016. https://commons.wikimedia.org/wiki/File:Leupeptin.svg.

Which of the following statements is the MOST likely explanation for the role of leupeptin in the extraction process?

A

Leupeptin acts as a buffer. In its absence, the change in pH upon lysis of the liver cells inactivated the enzyme.

B

Leupeptin acts as a protease inhibitor. In its absence, the enzyme was inactivated by natural proteases released from the liver cells.

C

Leupeptin acts as a compatible solute or osmolyte to stabilize the osmotic pressure of the solution and hence the liver cells. In its absence, the enzyme was inactivated by the change in osmotic pressure.

D

Leupeptin acts as a temperature stabilizer. In its absence, the enzyme was inactivated due to excessive heat generated during the extraction.

E

Leupeptin acts as an antioxidant. In its absence, the enzyme was inactivated by reactive free radicals produced when the cells were broken open during the extraction.

F

Leupeptin acts as a membrane permeabilizer. In its absence, the liver cells were not broken open and no enzyme was released into solution.