Question

Why does a sample of BSA treated with a reducing sample buffer display a different molecular weight than a sample of BSA treated with a nonreducing sample dilution buffer?

Answer 1

Ans. BSA is constituted of a single polypeptide chains with 17 intra-strand disulfide bonds.

Presence of reducing agents like b-mercaptoethanol in the reducing buffer cleaves all the disulfide bonds and roughly linearizes it. The protein thus moves through the get depending on its mass.

In a non-reducing buffer, all the disulfide bonds remain intact. The protein also retains its 3D structure to a large extent. The protein moves through the gel depending on it mass as well as 3D conformation.

Therefore, due to change in 3D conformation, BSA moves to different distances under reducing and non-reducing conditions causing the difference in its apparent molar mass under the two conditions.