Large-Scale Extraction of Recombinant Proteins from Bacteria
Adapted from Proteins and Proteomics, by Richard J. Simpson. CSHL Press, Cold Spring Harbor, NY, USA, 2003.INTRODUCTION
Bacteria are particularly convenient for producing recombinant proteins for purification purposes. Suitable extraction methods for bacterial cells include sonication, glass bead milling, grinding with alumina or sand, high-pressure shearing using the French pressure cell (French Press), and lysozyme treatment. These procedures are applicable for preparing extracts from a variety of Gram-negative bacteria such as Escherichia coli and Klebsiella pneumoniae, and Gram-positive bacteria such as Bacillus subtilis. Disruption of bacterial cells by enzymatic means is commonly used because a relatively uniform treatment is obtained when cells are in suspension. This protocol describes the enzymatic disruption of E. coli.
