Practicals will be divided into 2 modules:
Module 1 : Protein Trapping.
In these practical lessons, we will see how to determine the protein/amphipol mass ratio required to maintain soluble an integral membrane protein in the absence of detergent. We will use as model membrane protein bacteriorhodopsin (BR) from Halobacterium salinarum.
Module 2 : Protein Folding.
In these practical lessons, we will address the folding of BR using amphipols. In its native fold, BR binds a cofactor (retinal) via a Schiff base. The holoprotein is colored purple. When BR is denaturated in SDS, the retinal is released in solution and the sample becomes yellow. Upon renaturation, retinal spontaneously rebinds to bacterio-opsin (BO). The color is therefore a good indicator of the state (folded or unfolded) of the protein.