2 - Techniques and Basic Operations in Membrane Protein Purification

Abstract

Differential extraction and precipitation of proteins according to their solubility properties constitute the first step in many isolation protocols for membrane proteins. Protein enrichment by these easily performed initial steps is essential for several reasons: early removal of contaminants facilitates large-scale purification, which would otherwise be restricted by the limited capacity for protein of the separation media used in later steps; the spectrum of useful separation techniques is narrow, and protein isolation attempts might end with only partially purified proteins; and even if there are adequate purification techniques available, membrane protein stability is a limiting factor, and the purification protocols is improved if short times and few delipidating purification steps are involved. This chapter describes differential extraction by using bovine heart mitochondria as an example. About 30% of all mitochondrial proteins are water-soluble proteins, and about 70% are membrane proteins.