title: Molecular architecture of the Saccharomyces cerevisiae activated spliceosome.
abstract: The activated spliceosome (Bact) is in a catalytically inactive state and is remodeled into a catalytically active machine by the RNA helicase Prp2, but the mechanism is unclear. Here, we describe a 3D electron cryomicroscopy structure of the Saccharomyces cerevisiae Bactcomplex at 5.8-angstrom resolution. Our model reveals that in Bact, the catalytic U2/U6 RNA-Prp8 ribonucleoprotein core is already established, and the 5' splice site (ss) is oriented for step 1 catalysis but occluded by protein. The first-step nucleophile-the branchsite adenosine-is sequestered within the Hsh155 HEAT domain and is held 50 angstroms away from the 5'ss. Our structure suggests that Prp2 adenosine triphosphatase-mediated remodeling leads to conformational changes in Hsh155's HEAT domain that liberate the first-step reactants for catalysis.
authors: Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors: Reinhard
authors: R
authors: Rauhut
authors: Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors: Patrizia
authors: P
authors: Fabrizio
authors: Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors: Olexandr
authors: O
authors: Dybkov
authors: Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors: Klaus
authors: K
authors: Hartmuth
authors: Research Group Macromolecular Crystallography, MPI for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany.
authors: Vladimir
authors: V
authors: Pena
authors: 3D Electron Cryomicroscopy Group, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors: Ashwin
authors: A
authors: Chari
authors: Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors: Vinay
authors: V
authors: Kumar
authors: Bioanalytical Mass Spectrometry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany. Bioanalytics Group, Institute for Clinical Chemistry, University Medical Center, Göttingen, Robert-Koch-Straße 40, D-37075 Göttingen, Germany.
authors: Chung-Tien
authors: CT
authors: Lee
authors: Bioanalytical Mass Spectrometry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany. Bioanalytics Group, Institute for Clinical Chemistry, University Medical Center, Göttingen, Robert-Koch-Straße 40, D-37075 Göttingen, Germany.
authors: Henning
authors: H
authors: Urlaub
authors: Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany. b.kastner@mpi-bpc.mpg.de hstark1@gwdg.de reinhard.luehrmann@mpi-bpc.mpg.de.
authors: Berthold
authors: B
authors: Kastner
authors: 3D Electron Cryomicroscopy Group, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany. Department of 3D Electron Cryomicroscopy, Georg-August Universität, Göttingen, Justus von-Liebig-Weg 11, D-37077 Germany. b.kastner@mpi-bpc.mpg.de hstark1@gwdg.de reinhard.luehrmann@mpi-bpc.mpg.de.
authors: Holger
authors: H
authors: Stark
authors: Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany. b.kastner@mpi-bpc.mpg.de hstark1@gwdg.de reinhard.luehrmann@mpi-bpc.mpg.de.
authors: Reinhard
authors: R
authors: Lührmann
mesh_terms: D000251:Adenosine Triphosphatases; D055162:Biocatalysis; D020134:Catalytic Domain; D020285:Cryoelectron Microscopy; D005091:Exons; D011487:Protein Conformation; D020365:RNA Helicases / Q000737:chemistry / Q000235:genetics; D022821:RNA Splice Sites; D012326:RNA Splicing; D012342:RNA, Small Nuclear / Q000737:chemistry*; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000737:chemistry* / Q000235:genetics; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000737:chemistry* / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000648:ultrastructure*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics; D017381:Spliceosomes / Q000737:chemistry / Q000648:ultrastructure*
chemical_list: C077018:PRP8 protein, S cerevisiae; D022821:RNA Splice Sites; D012342:RNA, Small Nuclear; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C405484:U2 small nuclear RNA; D000251:Adenosine Triphosphatases; C104158:BRR2 protein, S cerevisiae; D020365:RNA Helicases