Which chromatographic technique is most appropriate to separate native carbonic anhydrase from its modified form?

• A. Anion-exchange chromatography
• B. Cation-exchange chromatography
• C. Gas-liquid chromatography
• D. Size-exclusion chromatography

Answer: (A)

To determine the most appropriate chromatographic technique for separating native carbonic anhydrase from its modified form, it is essential to understand the nature of the modifications that may occur. If the modified form of carbonic anhydrase carries a different charge than the native form, anion-exchange chromatography would be effective. This technique exploits differences in the net charge of proteins at a given pH.

Anion-exchange chromatography is best used when the target protein and its modified counterpart have different charges due to post-translational modifications, such as phosphorylation or glycosylation. By adjusting the pH of the mobile phase, you can promote the binding of one form over the other, allowing for effective separation.

In contrast, size-exclusion chromatography relies on the size of the molecules and would not effectively separate proteins that differ primarily in their charge. Cation-exchange chromatography would be appropriate only if the modifications result in a change that makes the modified form more positively charged compared to the native form. Gas-liquid chromatography is typically used for volatile substances and is not suitable for proteins.

Therefore, if the key difference between the native and modified forms of carbonic anhydrase is related to charge, anion-exchange chromatography is rightly identified as the most appropriate