Question

Two proteins are similar in size but differ significantly in the number of acidic and basic amino acids. Which of the following techniques would be best suited to separating these two proteins? (2 pts)

A. SDS-PAGE and gel filtration chromatography

B. Isoelectric focusing and dialysis

C. Immunoprecipitation and affinity chromatography

D. Isoelectric focusing and ion-exchange chromatography

E. None of the Above

*If possible please explain what molecules would corresond with each technique pairing in addition to what is the correct answer. My book does not explain the techniques very well and how they apply to molecule characteristics (size, charge, binding affinity, etc. )

Answer 1

Best method for separating two protein of same mass is isoelectric focusing and Ion exchange chromatography.

Isoelectric focusing is used to determine the affinity of protein for Acid and Base. ion exchange chromatography method- this technique is based on the net charge and pH of expose amino group and acid group of amino acid. In this process electroforesis performed on first positive charge column. When protein flow on positive charge column. All the negatively charged protein adhere with the column while neutral and positively charged protein flow out. After that wash the column with high concentration of salt two separate desired protein. Similar process can be apply for positively charged protein with negatively charged adsorption column.