Preparation of soluble proteins from Escherichia coli.
Purification of human IL-1beta is used in this unit as an example of the preparation of soluble proteins from E. coli. Bacteria containing IL-1beta are lysed, and IL-1 beta in the resulting supernatant is purified by anion-exchange chromatography, salt precipitation and cation-exchange chromatography, and then concentrated. Finally, the IL-1 beta protein is applied to a gel-filtration column to separate it from remaining higher- and lower-molecular-weight contaminants, the purified protein is stored frozen or is lyophilized. The purification protocol described is typical for a protein that is expressed in fairly high abundance (i.e., >5% total protein) and accumulates in a soluble state.
Wingfield
Paul T
PT
National Institutes of Health, Bethesda, Maryland.
D002411:Cation Exchange Resins; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification*; D012995:Solubility
D002411:Cation Exchange Resins; D011994:Recombinant Proteins