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; however, if a secretion vector is used, they can form in the periplasmic space. Inclusion bodies can be recovered from cell lysates by low-speed centrifugation. Following pre-extaction (or washing), protein is extracted from washed pellets using guanidine⋅HCl. The solubilized and unfolded protein is either directly folded or further purified by gel filtration in the presence of guanidine⋅HCl as described in this unit. A support protocol describes the removal of guanidine⋅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: D003593:Cytoplasm / Q000737:chemistry; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000302:isolation & purification*; D005822:Genetic Vectors; D002479:Inclusion Bodies / Q000737:chemistry* / Q000378:metabolism; D019897:Periplasm / Q000737:chemistry; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D012995:Solubility
chemical_list: D029968:Escherichia coli Proteins; D011994:Recombinant Proteins