# DBT JRF 2020 — Question 99

Exam: DBT JRF 2020 · Section: B · Topic: Biochemistry · Difficulty: Hard

## Question

Proteins are commonly purified by ion exchange chromatography (IEC) as a final step. Which of the following statements is NOT true?

## Options

- **A.** even proteins of similar isoelectric point can be conveniently separated by IEC, because interaction with the support is determined by the surface charge distribution of the protein rather than the net charge.
- **B.** in general, proteins can be eluted by increasing ionic strength
- **C.** above the isoelectric point, the proteins bind to anion exchangers.
- **D.** above the isoelectric point, the proteins bind to cation exchangers

## Answer

Option D

## Explanation

Ion exchange chromatography (IEC) separates proteins based on surface charge: ANION exchangers (positively charged resin, e.g. DEAE-cellulose, Q-Sepharose) bind negatively charged proteins. CATION exchangers (negatively charged resin, e.g. CM-cellulose, SP-Sepharose) bind positively charged proteins. At pH ABOVE the isoelectric point (pI): the net charge of the protein becomes NEGATIVE (protein has lost protons from basic groups and gained negative charges on acidic groups). Negatively charged proteins bind ANION exchangers. They would NOT bind cation exchangers (both are negatively charged — like charges repel). Therefore the statement 'above the isoelectric point, the proteins bind to cation exchangers' is FALSE. Elution is achieved by increasing ionic strength (NaCl gradient — salt ions compete for the binding sites). The surface charge distribution comment is true: proteins with similar pI can be separated by IEC because local charge patches matter.

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