Electrophoresis of a purified protein in SDS-PAGE in the presence of b-marcaptoethanol yields two bands of 35
kDa and 45 kDa. However, in a gel filtration chromatography, the same protein elutes as 80 kDa. What conclusion
can be drawn from the above observation?
(A) Two bands generated in SDS-PAGE due to degradation
(B) Protein is not purified to homogeneity

(C) Protein is a heterodimer

(D) Protein is a multimer

Correct answer: (C) Protein is a heterodimer

Explanation:

Let’s analyze the given data:

1. SDS-PAGE (with β-mercaptoethanol) yields two bands of 35 kDa and 45 kDa

   • SDS-PAGE separates proteins based on their molecular weight by denaturing them.
   • β-mercaptoethanol (BME) is a reducing agent that breaks disulfide bonds.
   • The presence of two distinct bands (35 kDa and 45 kDa) suggests that the protein consists of two different subunits.

2. Gel filtration chromatography elutes the protein as an 80 kDa complex

   • Gel filtration chromatography separates proteins based on size in their native, folded state.
   • The fact that the protein elutes as 80 kDa suggests that, in its native state, the protein is a complex of both subunits (35 kDa + 45 kDa).
   • This strongly indicates a heterodimer (a protein composed of two different subunits).

Why Not the Other Options?

• (A) Two bands generated in SDS-PAGE due to degradation ❌

  • If the protein was degraded, we would expect smaller, irregular fragments rather than two specific bands of 35 kDa and 45 kDa.

• (B) Protein is not purified to homogeneity ❌

  • If the protein was contaminated, SDS-PAGE would show multiple bands beyond just 35 kDa and 45 kDa.
  • The gel filtration chromatography result (80 kDa) matches the expected sum of these two subunits, supporting that this is a single, defined protein.

• (D) Protein is a multimer ❌

  • A multimer usually refers to a protein composed of more than two subunits (e.g., trimers, tetramers).
  • Here, the protein consists of only two different subunits, which is the definition of a heterodimer.

Conclusion:

The protein behaves as an 80 kDa complex in its native state and separates into two subunits (35 kDa and 45 kDa) in SDS-PAGE with a reducing agent, meaning it is a heterodimer.

✅ Correct answer: (C) Protein is a heterodimer.