Hydrophobic interaction chromatography (HIC) purifies proteins by leveraging their surface hydrophobicity, making it a key technique in biochemistry. The correct answer to the query is B. Hydrophobic amino acids on the protein surface.

Question Breakdown

This multiple-choice question tests core principles of protein purification methods used in molecular biology and competitive exams like GATE Life Sciences.

Option Analysis

• A. Charged amino acids: Incorrect, as charged amino acids (e.g., lysine, aspartate) drive ionic interactions in ion-exchange chromatography, not HIC.

• B. Hydrophobic amino acids on the protein surface: Correct. HIC exploits hydrophobic patches from amino acids like leucine, valine, or phenylalanine on the protein surface, enhanced by high salt buffers that promote binding to hydrophobic resins.

• C. Molecular weight: Incorrect; molecular weight is separated via size-exclusion chromatography, not hydrophobicity.

• D. Enzyme activity: Incorrect; activity-based purification uses affinity chromatography with substrates or inhibitors.

How HIC Works

Proteins bind to a hydrophobic stationary phase (e.g., phenyl or butyl resins) in high-salt conditions like 1-3 M ammonium sulfate, where salt reduces water around hydrophobic regions, strengthening interactions. Elution occurs by decreasing salt, releasing less hydrophobic proteins first.