title: Large-scale purification of ribosome-nascent chain complexes for biochemical and structural studies.
abstract: Here we present a method to purify large amounts of highly pure and stably arrested ribosome-nascent chain complexes (RNCs) from Escherichia coli cells. It relies on the combined use of translation-arrest sequences to generate nascent polypeptides of specified length and subsequent tag purification of the RNCs. Moreover, we adapted this method for the in vivo production of RNCs with specific isotope labeling of the nascent chains for nuclear magnetic resonance (NMR) studies. This method opens therefore possibilities for a wide range of biochemical and structural studies exploring conformations of nascent chains during the early steps of protein folding and targeting.
authors: Zentrum für Molekulare Biologie der Universität Heidelberg, DKFZ-ZMBH Alliance, Im Neuenheimer Feld 282, 69120 Heidelberg, Germany.
authors: Anna
authors: A
authors: Rutkowska
authors: Monika
authors: M
authors: Beerbaum
authors: Nandhakishore
authors: N
authors: Rajagopalan
authors: Jocelyne
authors: J
authors: Fiaux
authors: Peter
authors: P
authors: Schmieder
authors: Günter
authors: G
authors: Kramer
authors: Hartmut
authors: H
authors: Oschkinat
authors: Bernd
authors: B
authors: Bukau
mesh_terms: D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D010455:Peptides / Q000302:isolation & purification*; D014176:Protein Biosynthesis; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000737:chemistry* / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D046911:Macromolecular Substances; D010455:Peptides; D011993:Recombinant Fusion Proteins