title: Preparation and extraction of insoluble (inclusion-body) proteins from Escherichia coli.
abstract: High-level expression of many recombinant proteins in Escherichia coli leads to the formation of highly aggregated protein commonly referred to as inclusion bodies. Inclusion bodies are normally formed in the cytoplasm; alternatively, if a secretion vector is used, they can form in the periplasmic space. Inclusion bodies can be recovered from cell lysates and this unit describes preparation of washed pellets and solubilization of the protein using guanidine x HCl. The extracted protein, which is unfolded, is either directly folded as described in UNIT or further purified by gel filtration in the presence of guanidine x HCl as idescribed here. A support protocol describes the removal of guanidine x HCl from column fractions so they can be monitored by SDS-PAGE.
authors.affiliation: National Institutes of Health, Bethesda, Maryland.
authors.forename: Ira
authors.initials: I
authors.lastname: Palmer
authors.affiliation: National Institutes of Health, Bethesda, Maryland.
authors.forename: Paul T
authors.initials: PT
authors.lastname: Wingfield
mesh_terms: D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D019791:Guanidine / Q000737:chemistry; D011994:Recombinant Proteins / Q000235:genetics* / Q000302:isolation & purification*; D012995:Solubility; D014508:Urea / Q000737:chemistry
chemical_list: D011994:Recombinant Proteins; D014508:Urea; D019791:Guanidine