title: Wheat-germ cell-free production of prion proteins for solid-state NMR structural studies.
abstract: The expression of soluble, functional protein on a preparative scale poses a central challenge for structural studies. Cell-free protein expression has become a valuable alternative to cell-based methods, and allows today the expression of milligram quantities of protein. Its use is particularly attractive for NMR studies as it allows a multitude of isotopic labeling schemes. We have implemented and further developed protocols to prepare cell-free extracts from wheat germ to produce recombinant protein for solid-state NMR studies. Furthermore, we established the Renilla luciferase model to optimise and evaluate extract quality, and report first productions of the prions Ure2p and HET-s devoted to structural studies currently ongoing in our laboratories.
authors.affiliation: Institut de Biologie et Chimie des Protéines, UMR 5086 CNRS/Université de Lyon 1, 7 passage du Vercors, 69367 Lyon, France.
authors.forename: Claire
authors.initials: C
authors.lastname: Noirot
authors.forename: Birgit
authors.initials: B
authors.lastname: Habenstein
authors.forename: Luc
authors.initials: L
authors.lastname: Bousset
authors.forename: Ronald
authors.initials: R
authors.lastname: Melki
authors.forename: Beat H
authors.initials: BH
authors.lastname: Meier
authors.forename: Yaeta
authors.initials: Y
authors.lastname: Endo
authors.forename: François
authors.initials: F
authors.lastname: Penin
authors.forename: Anja
authors.initials: A
authors.lastname: Böckmann
mesh_terms: D000818:Animals; D002474:Cell-Free System / Q000378:metabolism*; D007553:Isotope Labeling; D049410:Luciferases, Renilla / Q000235:genetics / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011328:Prions / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D014908:Triticum / Q000196:embryology*
chemical_list: D011328:Prions; D011994:Recombinant Proteins; D049410:Luciferases, Renilla