Question

Which of the following methods can be used to detect a protein band in an organelle fraction for which there is no enzyme assay? A monoclonal antibody specific for the protein is available and each fraction obtained from the fractionation experiment will be assayed.

	a.	Immunoelectron microscopy
	b.	ELISA assay
	c.	Western blotting
	d.	Immunofluorescence

A membrane spanning protein that is a channel for potassium ion transport is made up of six copies of a single polypeptide. Potassium ions are transported across the membrane using the channel. In your experiments with the gene encoding the channel proteins, you have isolated a mutant of the potassium channel that transports very small amounts of potassium compared to the wild type. You perform SDS-PAGE electrophoresis with proteins from the mutant and proteins from the wild type. In the Coomassie blue stained gel, the lane for the wild type protein contains a protein band of 240 kDa and the lane for the mutant protein contains a protein band of 80 kDa. We can conclude that each mutant protein is______________________________the wild type subunits and that the molecular weight of each subunit in the mutant protein is ______________________________.

	a.	Shorter than, different
	b.	The same length, the same
	c.	There is not enough information to make a conclusion.
	d.	Shorter than, the same

What is true about the the rotation about bonds in a protein backbone?

	a.	All bonds in the backbone have restricted rotation and partial double-bond character.
	b.	The bond between the carbonyl carbon and nitrogen is restricted. Other bonds are free to rotate depending only on steric hindrance or the presence of proline residues.
	c.	The rotation is free about all bonds in the backbone, except for the bond between the nitrogen and alpha carbon in proline residues.
	d.	The rotation is free only about the peptide bond. The other bonds are restricted by steric hindrance and the presence of proline residues.

Answer 1

Question 1: Answer is option d. Immunofluorescence

In immunofluorescence, antibody is conjugated with fluorescence probe and this antibody is used for identification of protein of interest in the cell. Immunofluorescence is independent of enzymatic reactions.

Question 2: Answer is option a.

The mutant protein is shorter and has lesser molecular weight. The wildtype protein has 6 polypeptide chains. In SDS-PAGE there is a single band of 240kD. This implies that the molecular weight of each polypeptide is 240kD. Whereas in mutant protein, each polypeptide is 80kD.

Question 3: Answer is option b.

The bond between carbonyl carbon and nitrogen has a partial double bond character. Thus, all of the atoms linked to this bond are in a single plain. This bond is unrestricted for rotation.