Methods of reconstitution to investigate membrane protein function.

Membrane proteins are notoriously difficult to investigate in isolation. The focus of this chapter is the key step following extraction and purification of membrane proteins; namely reconstitution. The process of reconstitution re-inserts proteins into a lipid bilayer that partly resembles their native environment. This native environment is vital to the stability of membrane proteins, ensuring that they undergo vital conformational transitions and maintain optimal interaction with their substrates. Reconstitution may take many forms and these have been classified into two broad categories. Symmetric systems enable unfettered access to both sides of a bilayer. Compartment containing systems contain a lumen and are ideally suited to measurement of transport processes. The investigator is encouraged to ascertain what aspects of protein function will be undertaken and to apply the most advantageous reconstitution system or systems. It is important to note that the process of reconstitution is not subject to defined protocols and requires empirical optimisation to specific targets.