Molecular architecture of the Saccharomyces cerevisiae activated spliceosome

Reinhard Rauhut; Patrizia Fabrizio; Olexandr Dybkov; Klaus Hartmuth; Vladimir Pena; Ashwin Chari; Vinay Kumar; Chung-Tien Lee; Henning Urlaub; Berthold Kastner; Holger Stark; Reinhard Lührmann

doi:10.1126/science.aag1906

Molecular architecture of the Saccharomyces cerevisiae activated spliceosome

Science. 2016 Sep 23;353(6306):1399-1405. doi: 10.1126/science.aag1906. Epub 2016 Aug 25.

Authors

Affiliations

¹ Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
² Research Group Macromolecular Crystallography, MPI for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany.
³ 3D Electron Cryomicroscopy Group, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
⁴ 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.
⁵ Department of Cellular Biochemistry, Max Planck Institute (MPI) for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany. [email protected] [email protected] [email protected].
⁶ 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. [email protected] [email protected] [email protected].

PMID: 27562955
DOI: 10.1126/science.aag1906

Abstract

The activated spliceosome (B^act) 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 B^act complex at 5.8-angstrom resolution. Our model reveals that in B^act, 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.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adenosine Triphosphatases
Biocatalysis
Catalytic Domain
Cryoelectron Microscopy
Exons
Protein Conformation
RNA Helicases / chemistry
RNA Helicases / genetics
RNA Splice Sites
RNA Splicing
RNA, Small Nuclear / chemistry*
Ribonucleoprotein, U4-U6 Small Nuclear / chemistry*
Ribonucleoprotein, U4-U6 Small Nuclear / genetics
Ribonucleoprotein, U5 Small Nuclear / chemistry*
Ribonucleoprotein, U5 Small Nuclear / genetics
Saccharomyces cerevisiae / genetics
Saccharomyces cerevisiae / ultrastructure*
Saccharomyces cerevisiae Proteins / chemistry*
Saccharomyces cerevisiae Proteins / genetics
Spliceosomes / chemistry
Spliceosomes / ultrastructure*

Substances

PRP8 protein, S cerevisiae
RNA Splice Sites
RNA, Small Nuclear
Ribonucleoprotein, U4-U6 Small Nuclear
Ribonucleoprotein, U5 Small Nuclear
Saccharomyces cerevisiae Proteins
U2 small nuclear RNA
Adenosine Triphosphatases
BRR2 protein, S cerevisiae
RNA Helicases