Answer A1 is wrong This Table shows assay data from a purification procedure. Wh

Question

Answer A1 is wrong

This Table shows assay data from a purification procedure. Which statement is most likely TRUE? A. The gel-filtration step increased the specific activity but not the yield. A. Of the three chromatography steps, the affinity column step had the biggest drop in yield. A. After the CM-cellulose step, the purification factor reached over 1000, indicating effective purification. A. Most of the protein removed in the gel-filtration step was likely enzyme that had unfolded during the CM-cellulose step. A. All of the above are false.

Explanation / Answer

This data in this above explained table has shown that the yield from a purification step as well as yield. There is the quantity of it and then there is the activity. If we look into this carefully we can find out that the first step is a much generalized step where we get all the proteins in a mixture. Total activity is also more, next step is starting with the chromatography purification steps. Affinity column fraction keeps the activity in high standards while the non specific proteins are separated out. Another step followed by that is CM-cellulose fraction, is used for preparative separation of phospholipid classes. Here the separation step from has brought the yields to a narrowed down level which is the specific protein separation but the total activity is very high. Still the final step with Gel Filtration does the specific protein separation work which keep the activity low as the desired protein function and activity.

Now looking into the options given from above, the first one A1. HAs a decreased specific activity as there is only the desired protein with others non intended parts separated.

A2. This step from the Affinity chromatography is done to separate the proteins or macromolecules from the mixture, they are there to bind with the ligand. So from the cell fraction from centrifugation this steps has done the retention work of the protein molecules. Here the drop in yield is the highest most, so this option id chosen to be right.

A3. Purification factor is to be calculated by this formula specific activity of (x) __= purification factor for (x) specific activity of starting material. For each fraction, divide the enzyme concentration by the protein concentration. If we do the calculation it does not come to 1000 as it is mentioned in the comment. Specific activity of CM-cellulose affect is 330000/600 = 550, and Specific activity from starting material is 350000/1000 = 350, now dividing both, 550/350 = 1.57 units.

A4. In CM cellulose step is where ion exchange takes place, as some protein and enzymes molecules re bigger in size so they are separated out by the CM cellulose exchange. Here the filtration separates the proteins out of the system and only the specific protein having relevant ions can get exchanged and elute out either with our interest or they remain in the matrix and later separated out.

The interesting fact about the gel filtration part is this solely based on the separation of proteins, peptides, and oligonucleotides on the basis of size. Molecules move through a bed of porous beads, diffusing into the beads and the smaller molecules diffuse further into the pores, so as the option suggests that the bigger molecules are likely to be separated as they are unfolded during the CM-cellulose exchange step may be right or may not be. Because the CM cellulose step does not suggest the macromolecules to be opened up.

Conclusion, A2 answer choice is likely to be true.

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