Question

Which of the following chromatography separates according to the sign and magnitude of the net electric charges of the proteins at a given pH?

• A. Size exclusion chromatography
• B. Affinity chromatography
• C. Isoelectric focusing
• D. Ion-exchange chromatography

Hint:

Remember:
• Ion-exchange chromatography \(\rightarrow\) Charge-based separation
• Size exclusion chromatography \(\rightarrow\) Size-based separation
• Affinity chromatography \(\rightarrow\) Specific binding interactions
• Isoelectric focusing \(\rightarrow\) Separation by isoelectric point (\(pI\))

Answer 1

The Correct Option is D

Concept: Ion-exchange chromatography separates proteins based on: \[ \text{Net electric charge of proteins at a given pH} \] Proteins carry positive or negative charges depending on the pH of the surrounding medium relative to their isoelectric point (\(pI\)). In ion-exchange chromatography:
• Positively charged proteins bind to cation exchangers
• Negatively charged proteins bind to anion exchangers Thus, separation depends on both:
• Sign of charge
• Magnitude of charge

Step 1: Identifying the chromatography based on protein charge. The question specifically mentions: \[ \text{Separation according to sign and magnitude of net electric charge} \] This is the principle of: \[ \boxed{\text{Ion-exchange chromatography}} \] Proteins interact with oppositely charged groups attached to the chromatography matrix.

Step 2: Understanding the other chromatography methods.
• Option (A): Size exclusion chromatography separates molecules based on size and shape.
• Option (B): Affinity chromatography separates molecules based on specific biological interactions such as antigen-antibody or enzyme-substrate binding.
• Option (C): Isoelectric focusing separates proteins according to their isoelectric point (\(pI\)), not directly by magnitude of net charge. Hence, the correct answer is ion-exchange chromatography. \[ \boxed{\text{Protein charge separation} \Rightarrow \text{Ion-exchange chromatography}} \]