Q: Which equation best describes the relationship between the net charge of a protein and its elution from an anion-exchange column?

$$\text{Elution order} \propto |Q_{net}|$$

Question 1

Which stationary phase is typically used in anion exchange chromatography to purify proteins with a net negative charge at physiological pH?

Accepted Answer

Diethylaminoethyl (DEAE) cellulose

Question 2

A peptide with the sequence Gly-Ala-Asp-Glu-Ser is loaded onto an anion-exchange column at pH 7.0. Which property will most influence its elution order compared to a peptide with the sequence Gly-Ala-Lys-Arg?

Accepted Answer

The net negative charge of the peptide

Question 3

If a protein has a pI (isoelectric point) of 5.0 and is loaded onto an anion-exchange column at pH 8.0, what will be its net charge and how will it interact with the column?

Accepted Answer

It will have a net negative charge and bind to the column

Question 4

Which amino acid would elute first from an anion-exchange column at physiological pH?

Accepted Answer

Alanine

Question 5

A mixture of histidine (pI = 7.6) and arginine (pI = 10.8) is separated by ion-exchange chromatography at pH 8.0. Which type of column should be used for optimal separation?

Accepted Answer

Cation-exchange chromatography at pH 8

Question 6

Given the following peptide: Tyr-Ala-Asp-Glu-Ser, which residue is most responsible for its retention on an anion-exchange column at pH 7.0?

Accepted Answer

Aspartate

Question 7

Which equation best describes the relationship between the net charge of a protein and its elution from an anion-exchange column?

Accepted Answer

$$	ext{Elution order} \propto |Q_{net}|$$

Question 8

A protein sample is loaded onto a DEAE-cellulose column at pH 7.5. Which of the following proteins will elute last?

Accepted Answer

A protein with pI = 4.5

Question 9

Which of the following statements about ion-exchange chromatography is correct?

Accepted Answer

Anion-exchange columns bind negatively charged proteins

Question 10

A peptide containing the sequence Asp-His-Glu is loaded onto an anion-exchange column at pH 9.3. Which amino acid will most strongly interact with the column?

Accepted Answer

Aspartate

Question 11

If a protein does not bind to a DEAE-cellulose column at pH 7.0, what can you infer about its isoelectric point (pI)?

Accepted Answer

Its pI is close to or above 7.0

Question 12

Which stationary phase would you use to purify a protein with a net positive charge at pH 7.0?

Accepted Answer

Carboxymethyl (CM) cellulose

Question 13

A mixture of peptides is loaded onto an anion-exchange column at pH 9.3. Which peptide will elute first?

Accepted Answer

A peptide with the most positive net charge

Question 14

Which of the following best explains why increasing the salt concentration in the elution buffer causes proteins to elute from an ion-exchange column?

Accepted Answer

Salt ions compete with protein charges for binding sites on the column

Question 15

A protein with a pI of 6.0 is loaded onto a DEAE-cellulose column at pH 8.0. What is the expected net charge of the protein and its behavior?

Accepted Answer

Net negative charge; binds to the column

Question 16

Which of the following amino acids would NOT bind to an anion-exchange column at pH 7.0?

Accepted Answer

Alanine

Question 17

Which of the following is the correct formula for calculating the net charge of a peptide at a given pH?

Accepted Answer

$$Q_{net} = \sum_{i} q_{i}$$, where $$q_{i} is $$the charge of each ionizable group at the given pH

Question 18

A peptide with the sequence Gly-Ala-Lys-Arg is loaded onto an anion-exchange column at pH 7.0. What is the expected outcome?

Accepted Answer

It will elute quickly because it has a net positive charge

Question 19

Which of the following is a common misconception about anion-exchange chromatography?

Accepted Answer

Anion-exchange columns bind positively charged proteins

Anion Exchange Chromatography: Principles and Applications in Protein Purification

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