The GST-Glutathione is an example of ______ affinity chromatography (a)Antibody-antigen (b)Dye-affinity (c)Enzyme-substrate (d)Metal-affinity

The GST-Glutathione is an example of ______ affinity chromatography

Glutathione is a molecule that has a role inAnswers: A. amino acid oxidationB. protection against reactive oxygen speciesC. glycolysisD. TCA cycle

Human gamma-glutamyl transpeptidase (hGGT) is an enzyme involved in cysteine homeostasis, and its overexpression has been linked to asthma, cancer, and other diseases.  It catalyzes the addition of water across a bond to cleave extracellular glutathione (GSH) into glutamate and a dipeptide composed of cysteine and glycine.  It is believed to function by the mechanism shown in Figure 1.Figure 1  Proposed mechanism of hGGT-mediated GSH cleavageResearchers interested in investigating hGGT inhibitors as a treatment for pathologies related to excess hGGT activity performed the following experiments, each containing the same concentration of hGGT.Experiment 1hGGT was incubated with various concentrations of GSH in the presence or absence of 250 μM candidate compound acivicin or Compound 1.  However, both candidates were found to be unsuitable as acivicin showed significant neurotoxicity and Compound 1 acted as an activator instead of an inhibitor.Figure 2  Lineweaver-Burk plot of hGGT activity with respect to GSH in the presence or absence of 250 μM candidate inhibitorsExperiment 2Researchers hypothesized that slight variations in the structure of Compound 1 could result in hGGT inhibition and tested several derivatives, shown in Table 1.Table 1  Kinetic Parameters of hGGT in the Presence or Absence of Several Candidate Inhibitors at 250 μMExperiment 3To better understand the physiological conditions under which hGGT operates, researchers exposed a cultured cell line expressing hGGT on the cell surface to a solution that mimicked the composition of extracellular fluid, including levels of GSH.  They found that in this setup, uninhibited hGGT results in a reaction rate that is approximately ½Vmax.Adapted from Wickham, S., Regan, N., West, M.B., Thai, J., Cook, P.F., Terzyan, S.S., Li, P.K., & Hanigan, M.H. (2013). Inhibition of human ?-glutamyl transpeptidase: development of more potent, physiologically relevant, uncompetitive inhibitors. The Biochemical Journal, 450(3), 547–557. Question 24Based on the information in Experiment 3, if researchers wish to further study the inhibitor that is most effective under physiological conditions, they should choose the one that yields the lowest hGGT activity when GSH concentration is:A.orders of magnitude less than the Km of the uninhibited enzyme.B.approximately equal to the Km of the uninhibited enzyme.C.approximately four times the Km of the uninhibited enzyme.D.orders of magnitude greater than the Km of the uninhibited enzyme.

What can be seen in this picture?Question 3AnswerA.the column used in affinity chromatographyB.the bilayer prepared during in vitro translationC.agarose gel prepared for gel electrophoresisD.cuvette with sample, to measure the protein concentrationE.sample prepared for analysis by the Biuret method

Match the stationary phase (Column I) with its corresponding chromatographytechnique (Column II).Column I Column IIP. Protein A 1. Size exclusion chromatographyQ. Sephadex 2. Ion-exchange chromatographyR. Phenylsepharose 3. Affinity chromatographyS. Diethylaminoethyl cellulose 4. Hydrophobic interaction chromatography(A) P-1, Q-4, R-2, S-3(B) P-3, Q-1, R-4, S-2(C) P-3, Q-4, R-2, S-1(D) P-4, Q-1, R-3, S-2