Multiple Choice
Which amino acid elutes last from an anion-exchange column at physiological pH?
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5. Protein Techniques
Anion-Exchange Chromatography
Multiple Choice
Which type of ion exchange chromatography would be best to separate a mixture of histidine and arginine?
His: pK1 = 1.8, pK2 = 9.3, pKR = 6.0 Arg: pK1 = 1.8, pK2 = 9.0, pKR = 12.5
A
Anion-exchange chromatography at pH = 2.
B
Anion-exchange chromatography at pH = 4.
C
Cation-exchange chromatography at pH = 2.
D
Cation-exchange chromatography at pH = 4.
E
Cation-exchange chromatography at pH = 9.
Verified step by step guidance1
Understand the principle of ion exchange chromatography: It separates molecules based on their net charge at a given pH. Cation-exchange chromatography uses a negatively charged resin to bind positively charged molecules, while anion-exchange chromatography uses a positively charged resin to bind negatively charged molecules.
Determine the net charge of histidine and arginine at different pH levels: Use the pKa values provided to calculate the net charge of each amino acid at the specified pH levels. The net charge is determined by the ionization state of the amino acid's functional groups.
Calculate the isoelectric point (pI) for each amino acid: The pI is the pH at which the amino acid has no net charge. For histidine, the pI is calculated using the average of pK2 and pKR, and for arginine, it is the average of pK2 and pKR.
Evaluate the net charge of histidine and arginine at pH 9: At this pH, compare the net charges of histidine and arginine to determine which will be positively charged and thus bind to a cation-exchange resin.
Select the appropriate ion exchange method: Since both histidine and arginine will have different net charges at pH 9, cation-exchange chromatography is suitable for separating them based on their positive charges.
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