Defining Membrane Protein Localization by Isopycnic Density Gradients

Bacterial membrane proteins account for around one-third of the proteome in many species and can represent much more than half of the mass of the membranes. Classic techniques in cell biology can be applied to characterize bacterial membranes and their membrane protein constituents, and here we describe a protocol for the purification of outer membranes and inner membranes from Escherichia coli. This allows for compositional analysis of the membranes as well as functional analyses. The procedure can be applied with minor modifications to other bacterial species including those carrying capsular polysaccharide attached to the outer membrane.