Question

A protein was purified to homogeneity. The determination of the mass by gel‑filtration chromatography yields a value of 60 kDa. Chromatography in the presence of urea results in a 30 kDa species. Repeating the chromatography in the presence of both urea and β‑mercaptoethanol, however, results in a single molecular species of 15 kDa. What do the data suggest about the structure of the protein?

Answer 1

In the given case, the homogeneity of the protein is done. By performing, the technique of gel-filtration chromatography, the mass of the protein obtained is 60 kilodalton. When the chromatography is performed in the existence of urea, a protein of mass 30 kilodalton is obtained. The ratio between the two obtained mass is 60/30 = 2, thus, it shows that the protein comprises two subunits and each of them weighs 30 kilodaltons.

When the same technique of chromatography is performed in the existence of beta-mercaptoethanol and urea, a single protein of mass 15 kilodalton is produced. Now, the ratio is 30/15 = 2. Therefore, each subunit of the protein comprises two units of mass 15 kilodalton.

Hence, it shows that the protein is a tetramer of 15 kilodalton subunits and each subunit produces dimers of mass 30 kilodalton and is attached by disulfide bonds.