Fermentation and growth of Escherichia coli for optimal protein production.
Large-scale production of recombinant proteins in Escherichia coli requires growth of cells in fermentors. This unit lists E. coli strains appropriate for use in fermentors, and also discusses important characteristics of fermentation equipment. Production of recombinant proteins in batch fermentations is described, as are variations of fermentation systems that enable continuous growth and protein production in high-cell-density, fed-batch cultures and that permit labeling of recombinant proteins with heavy atom derivatives such as selenomethionine or with stable isotopes such as (2)H, (13)C, and (15)N. Production of labeled proteins facilitates structural studies by X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy. The protocols in this unit are designed for expression systems directing intracellular or periplasmic localization of recombinant proteins; however, in the case of extracellular secretion of the desired protein, the culture medium itself, rather than pelleted cells, would be saved, concentrated, and subjected to purification processes. Fermentation experiments require careful monitoring of cell growth and assurance of preinoculation sterility, both which are described here.
Bernard
A
A
Glaxo Institute for Molecular Biology, Geneva, Switzerland.
Payton
M
M
D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D005285:Fermentation; D009682:Magnetic Resonance Spectroscopy; D011506:Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
D011506:Proteins; D011994:Recombinant Proteins