Assessments for Animated Techniques

3.1 SDS-PAGE Analysis

Chapter 3. 3.1 SDS-PAGE Analysis

Assessments for Animated Techniques

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3.1 SDS-PAGE Analysis

1.

The purpose of adding SDS to the protein sample prior to loading the sample on the gel is to:

disrupt the disulfide bridges within the protein.

cause the proteins to migrate based on their electric charge (isoelectric point).

denature the protein and coat it with a negative charge.

cleave the protein into smaller fragments to facilitate protein sequencing.

_feedback_correct: Correct. _feedback_incorrect: Incorrect.

2.

The purpose of adding β-mercaptoethanol to the protein sample prior to loading the sample on the gel is to:

disrupt the disulfide bridges within the protein.

cause the proteins to migrate based on their electric charge (isoelectric point).

denature the protein and coat it with a negative charge.

cleave the protein into smaller fragments to facilitate protein sequencing.

_feedback_correct: Correct. _feedback_incorrect: Incorrect.

3.

Which of the following statements about gel electrophoresis is false?

Proteins in an SDS-PAGE gel run from the anode to the cathode when placed in an electric field.

Proteins are separated based on their isoelectric point.

Proteins are separated based on their molecular weight.

Charged dyes are often used to monitor the progress of gel electrophoresis.

_feedback_correct: Correct. _feedback_incorrect: Incorrect.

4.

After running gel chromatography on your purified protein sample, you notice that the molecular weight of the protein corresponds to 120 kDa.

a. When you run the same sample out using SDS-PAGE, you notice two bands, one at 50 kDa and one at 20 kDa. What can you conclude about the structure of your protein?

_feedback: The protein is a trimer composed of two 50 kDa subunits and one 20 kDa subunit.

b. If the same protein sample is treated with β-mercaptoethanol prior to running the SDS-PAGE gel, the result is one 50 kDa band, one 5 kDa band, and one 15 kDa band. What can you conclude about the structure of the protein?

_feedback: The 20 kDa subunit is really two peptide subunits, a 15 kDa subunit and a 5 kDa subunit, held together by a disulfide bridge.

5.

True or False: Larger proteins move more quickly through the gel than the smaller ones.

True

False

_feedback_correct: Correct. _feedback_incorrect: Incorrect.

6.

True or False: Glycerol is added to the loading buffer to make the protein sample heavier than the running buffer, ensuring that the sample will sink in the well when it is loaded.

True

False

_feedback_correct: Correct. _feedback_incorrect: Incorrect.

7.

Treating proteins with SDS and heat allows all of the proteins to have the same , allowing the proteins to be separated based on their size.

_feedback_correct: Correct. _feedback_incorrect: Incorrect.

8.

You have purified a protein to homogeneity and are running a sample on SDS-PAGE to visualize it. You notice that there are two bands on the gel that are nearly the same mass in kDa. Using an antibody and Western blotting, you confirm that both bands represent the protein that you are trying to purify. Treatment of the sample with β-mercaptoethanol does not change the appearance of either band. What is the most likely explanation for the appearance of two bands? How would you try to test this?

_feedback: Some of the protein may have a posttranslational modification such as phosphorylation or acetylation that has occurred. This could be tested for by treating the sample with a phosphatase or a deacetylase prior to SDS-PAGE analysis.

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Chapter 3. 3.1 SDS-PAGE Analysis

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