title: Current strategies for protein production and purification enabling membrane protein structural biology.
abstract: Membrane proteins are still heavily under-represented in the protein data bank (PDB), owing to multiple bottlenecks. The typical low abundance of membrane proteins in their natural hosts makes it necessary to overexpress these proteins either in heterologous systems or through in vitro translation/cell-free expression. Heterologous expression of proteins, in turn, leads to multiple obstacles, owing to the unpredictability of compatibility of the target protein for expression in a given host. The highly hydrophobic and (or) amphipathic nature of membrane proteins also leads to challenges in producing a homogeneous, stable, and pure sample for structural studies. Circumventing these hurdles has become possible through the introduction of novel protein production protocols; efficient protein isolation and sample preparation methods; and, improvement in hardware and software for structural characterization. Combined, these advances have made the past 10-15 years very exciting and eventful for the field of membrane protein structural biology, with an exponential growth in the number of solved membrane protein structures. In this review, we focus on both the advances and diversity of protein production and purification methods that have allowed this growth in structural knowledge of membrane proteins through X-ray crystallography, nuclear magnetic resonance (NMR) spectroscopy, and cryo-electron microscopy (cryo-EM).
authors.affiliation: a Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, NS B3H 4R2, Canada.
authors.forename: Aditya
authors.initials: A
authors.lastname: Pandey
authors.affiliation: a Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, NS B3H 4R2, Canada.
authors.forename: Kyungsoo
authors.initials: K
authors.lastname: Shin
authors.affiliation: a Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, NS B3H 4R2, Canada.
authors.forename: Robin E
authors.initials: RE
authors.lastname: Patterson
authors.affiliation: a Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, NS B3H 4R2, Canada.
authors.forename: Xiang-Qin
authors.initials: XQ
authors.lastname: Liu
authors.affiliation: a Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, NS B3H 4R2, Canada.|b Department of Chemistry, Dalhousie University, Halifax, NS B3H 4R2, Canada.
authors.forename: Jan K
authors.initials: JK
authors.lastname: Rainey
mesh_terms: D000818:Animals; D018360:Crystallography, X-Ray; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins
keywords: biologie structurale; enrichissement d’isotopes stables; expression et purification des protéines; fusion proteins; membrane protein; protein expression and purification; protéine membranaire; protéines de fusion; stable isotope enrichment; structural biology