What kind of ion-exchange column would you use to separate a mixture of the 2 pe

Question

What kind of ion-exchange column would you use to separate a mixture of the 2 peptides at pH 1? What would be the elution order? What kind of ion-exchange column would you use to separate a mixture of the 2 peptides at pH 13? What would be the elution order? The relevant pKa values for the ionizable groups are: Alpha amino group (at N-terminal): 8 Alpha carboxyl group (at C-terminal): 2 Glu and Asp side-chain carboxyl groups: 4 His side-chain: 6 Lys side-chain: 10 Arg side-chain: 12 Note that the carboxyl groups are potentially negatively charged dep on pH (neutral when protonated at low pH): the other groups are all

Explanation / Answer

10b) Cation exchange column(CEC) is used for seperation of peptides at pH 1. It uses negatively charged ion exchange resin with an affinity for molecule having net positive surface charge. Peptides are eluted by linear elution.

10c) Anion exchange column( AEC) is used for seperation of 2 peptides at pH 13. Anion exchange chromatography uses positively charged ion exchange resin with an affinity for molecule having net negative charge. A salt gradient is used to seperate the peptide from other bound to it. Peptides with pI(Isoelectric point) values closer to 8.5 will elute at lower ionic strength and peptides with very low pI values elute at high salt concentration. Peptides are eluted by step elution.

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