128. A laboratory is trying to purify a Histone-binding protein from its native source. Which one of
the following purification techniques should be used for one-step purification of the protein?
A. Cation exchange chromatography
B. Immuno affinity chromatography
C. Gel exclusion chromatography
D. Metal affinity chromatography

Introduction:

Purifying a histone-binding protein from its native cellular environment presents unique challenges due to the complexity of protein interactions and the presence of numerous similar molecules. For high specificity and efficiency in a single-step purification, selecting the correct chromatographic method is crucial. Among the available options, immunoaffinity chromatography stands out as the ideal technique.

Explanation of Purification Techniques:

Let’s examine the given methods:

A. Cation Exchange Chromatography

This method separates proteins based on net positive charge. While histone-binding proteins may carry a charge, this technique lacks the target specificity needed for one-step purification from a native lysate.

B. Immunoaffinity Chromatography (Correct Answer)

This technique uses antibodies specific to the target protein, enabling highly selective binding and purification. For histone-binding proteins, if a specific antibody is available, this method offers unmatched purity in a single step, directly from complex mixtures like nuclear extracts.

C. Gel Exclusion Chromatography (Size Exclusion)

This separates proteins based on molecular size. However, proteins with similar sizes will elute together, making this unsuitable for one-step purification in a native system.

D. Metal Affinity Chromatography

Typically used for recombinant proteins tagged with His-tags (e.g., His₆-tag). Native histone-binding proteins do not possess such tags naturally, making this ineffective unless the protein is expressed recombinantly.

Why Immunoaffinity Chromatography Is Ideal:

• High specificity due to antigen-antibody interaction

• Effective for low abundance proteins

• Can be used directly on native lysates

• Provides high purity in one step, eliminating the need for further rounds of purification

Correct Answer:

B. Immunoaffinity Chromatography

Conclusion:

For purifying a histone-binding protein from its native source, immunoaffinity chromatography is the best one-step purification method. It allows specific capture and elution of the target protein with minimal contamination, enabling fast and efficient downstream analysis