title: Membrane Protein Production in Escherichia coli: Protocols and Rules.
abstract: Functional and structural studies on membrane proteins are limited by the difficulty to produce them in large amount and in a functional state. In this review, we provide protocols to achieve high-level expression of membrane proteins in Escherichia coli. The T7 RNA polymerase-based expression system is presented in detail and protocols to assess and improve its efficiency are discussed. Protocols to isolate either membrane or inclusion bodies and to perform an initial qualitative test to assess the solubility of the recombinant protein are also included.
authors.affiliation: Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, Institut de Biologie Physico-Chimique, CNRS, Univ Paris Diderot, Sorbonne Paris Cité, PSL research university, Paris, France.
authors.forename: Federica
authors.initials: F
authors.lastname: Angius
authors.affiliation: Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, Institut de Biologie Physico-Chimique, CNRS, Univ Paris Diderot, Sorbonne Paris Cité, PSL research university, Paris, France.
authors.forename: Oana
authors.initials: O
authors.lastname: Ilioaia
authors.affiliation: Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, Institut de Biologie Physico-Chimique, CNRS, Univ Paris Diderot, Sorbonne Paris Cité, PSL research university, Paris, France.
authors.forename: Marc
authors.initials: M
authors.lastname: Uzan
authors.affiliation: Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, Institut de Biologie Physico-Chimique, CNRS, Univ Paris Diderot, Sorbonne Paris Cité, PSL research university, Paris, France. Bruno.Miroux@ibpc.fr.|Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR7099 CNRS IBPC, 13 rue Pierre et Marie Curie, 75005, Paris, France. Bruno.Miroux@ibpc.fr.
authors.forename: Bruno
authors.initials: B
authors.lastname: Miroux
mesh_terms: D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development*; D017408:Guidelines as Topic; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000096:biosynthesis; D012995:Solubility; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases
keywords: E. coli; Production of recombinant proteins; T7 RNA polymerase