Question

SDS Page Gel:

The provided standard protein sample for electrophoresis consists of 9 polypeptides with molecular weights ranging from 250 to 15 KDa.

Sample 1: Protein A in a sample buffer with B-Mercaptoethanol

Sample 2: Protein A in a sample buffer without B-Mercaptoethanol

Sample 3: Protein B in a sample buffer with B-Mercaptoethanol

Sample 4: Protein C in a sample buffer without B-Mercaptoethanol

Use the picture below & the information about the proteins above to answer the following questions.

1a. Fill out the table below based on the characteristics of the proteins and the components of the sample buffer

	Sample 1	Sample 2	Sample 3	Sample 4
Molecular Weight in the Gel (KDa)
Protein Oligomerization
Molecular Weight of Each Protein Subunit

1b. Explain for each sample how the B-mercaptoethanol is affecting the oligomerization state of the protein

1c, How would you confirm the oligimerization state of protein C (sample 4)? Why?

time 2. SDS page gel. (18 points) The provided standard protein sample (Dual color-Biorad) for electrophoresis consists of 9 polypeptides with molecular weights ranging from 250 to 15 KDa. Using the picture of this model gel, fill in the table below. All samples were boiled and have SDS in the sample buffer. STD 1 2 3 4 250 150 100 75

Answer 1

	Sample 1	Sample2	Sample 3	Sample 4
MW (Kda)	~58	~58	~60	~65
Protein oligomerization	Yes	No	Yes	No
MW of subunit	Single band	Single band	~60 and ~55 Kda (crude fraction)	Single band

1 b. Beta mercaptoethanol (2-ME) is a denaturing agent, which denature the protein and breakdown the disulfide bonds present in the polypeptide. When we store sample in deep freezer then they oligomerizeagain that's why every time when we want to see them on SDS-PAGE we boil our samples, so that 2-ME can act effectively.

1 c. 2-ME inhibits oligomerization, so in sample 4 there would be oligomerization but sample is a purified fraction that's why we can observe only one thick bands which further confirms the oligomerization of protein C.