Consider the mixture of proteins listed below for questions 10-12: You load the
ID: 486071 • Letter: C
Question
Consider the mixture of proteins listed below for questions 10-12: You load the protein mix onto a cation exchange column at pH 5. Next, you apply a "washing" step by passing through buffer at pH 5. Finally, for your elution step, you apply a pH gradient starting from pH 2.0 to pH 13.0 (A gradient buffer system allows you to gradually and continuously change the pH of your mobile phase starting from pH 2 up to pH 13). Indicate the order in which proteins will exit the column during the elution step. Please explain your answerExplanation / Answer
This technique of protein separation is based on pI (isoelectric point). pI of a protein is defined as the pH at which the protein is neutral. In a cationic exchanger column, the adsorbent is negatively charged. The protein with pI lesser than the pH will be negatively charged and elute. Thus when this seperation process is started, only A has pI less than pH of 5 and will be eluted first. The other proteins will have positive charge and will bind to the column. Protein B will be eluted 7 pH followed by D, E and finally C.
The order of protein elution is A, B, D, E, C.