Question

Proteins are separated on the basis of size by _______.

• A. SDS-PAGE
• B. HPLC
• C. Affinity chromatography
• D. Ion-exchange chromatography

Hint:

In SDS-PAGE, the detergent SDS denatures proteins and gives them a uniform negative charge-to-mass ratio. This ensures that migration through the polyacrylamide gel is determined solely by molecular weight (size).

Answer 1

The Correct Option is A

Step 1: Understanding the Question:
The question asks for the standard analytical laboratory technique used to separate a mixture of proteins based on their molecular weight (size).

Step 2: Detailed Explanation:

• SDS-PAGE (Sodium Dodecyl Sulfate - Polyacrylamide Gel Electrophoresis): This is a widely used biochemistry technique for separating proteins strictly by their molecular weight.

• Mechanism of SDS-PAGE:

• Denaturation and Charge Normalization: Proteins are treated with SDS, an anionic detergent. SDS denatures the tertiary structure of proteins and binds to them, imparting a uniform negative charge proportional to the protein's mass. This eliminates the influence of native shape and charge.

• Electrophoretic Separation: The negatively charged, unfolded proteins are loaded onto a polyacrylamide gel and exposed to an electric field. The proteins migrate toward the positive anode.

• Sieving Effect: The polyacrylamide gel acts as a molecular sieve. Small proteins move easily through the gel pores and migrate faster, while larger proteins are retarded and migrate slower. This separates them strictly by size.

• Why Other Options are Less Specific for Size:

• HPLC (High-Performance Liquid Chromatography): Separates molecules based on chemical polarity (reverse-phase), size, or ion charge, depending on the column chemistry, but is not primarily used for simple molecular weight sizing of whole protein mixtures.

• Affinity Chromatography: Separates proteins based on highly specific biological binding interactions (e.g., antigen-antibody or enzyme-substrate), not size.

• Ion-Exchange Chromatography: Separates proteins based on their net surface charge at a specific pH.

Step 3: Final Answer:
SDS-PAGE is the standard technique used to separate proteins based on their molecular size, making option (A) the correct choice.