title: Cryo-EM structure of a human spliceosome activated for step 2 of splicing.
abstract: Spliceosome rearrangements facilitated by RNA helicase PRP16 before catalytic step two of splicing are poorly understood. Here we report a 3D cryo-electron microscopy structure of the human spliceosomal C complex stalled directly after PRP16 action (C*). The architecture of the catalytic U2-U6 ribonucleoprotein (RNP) core of the human C* spliceosome is very similar to that of the yeast pre-Prp16 C complex. However, in C* the branched intron region is separated from the catalytic centre by approximately 20 Å, and its position close to the U6 small nuclear RNA ACAGA box is stabilized by interactions with the PRP8 RNase H-like and PRP17 WD40 domains. RNA helicase PRP22 is located about 100 Å from the catalytic centre, suggesting that it destabilizes the spliced mRNA after step two from a distance. Comparison of the structure of the yeast C and human C* complexes reveals numerous RNP rearrangements that are likely to be facilitated by PRP16, including a large-scale movement of the U2 small nuclear RNP.
authors.affiliation: Department of Structural Dynamics, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Karl
authors.initials: K
authors.lastname: Bertram
authors.affiliation: Department of Cellular Biochemistry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Dmitry E
authors.initials: DE
authors.lastname: Agafonov
authors.affiliation: Department of Structural Dynamics, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Wen-Ti
authors.initials: WT
authors.lastname: Liu
authors.affiliation: Department of Cellular Biochemistry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Olexandr
authors.initials: O
authors.lastname: Dybkov
authors.affiliation: Department of Cellular Biochemistry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Cindy L
authors.initials: CL
authors.lastname: Will
authors.affiliation: Department of Cellular Biochemistry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Hartmuth
authors.affiliation: Bioanalytical Mass Spectrometry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.|Bioanalytics Group, Institute for Clinical Chemistry, University Medical Centre Göttingen, Robert-Koch-Straße 40, D-37075 Göttingen, Germany.
authors.forename: Henning
authors.initials: H
authors.lastname: Urlaub
authors.affiliation: Department of Cellular Biochemistry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Berthold
authors.initials: B
authors.lastname: Kastner
authors.affiliation: Department of Structural Dynamics, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Holger
authors.initials: H
authors.lastname: Stark
authors.affiliation: Department of Cellular Biochemistry, MPI for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Lührmann
mesh_terms: D000241:Adenosine / Q000378:metabolism; D001483:Base Sequence; D055162:Biocatalysis; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D020285:Cryoelectron Microscopy; D053487:DEAD-box RNA Helicases / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D005091:Exons / Q000235:genetics; D006801:Humans; D007438:Introns / Q000235:genetics; D008958:Models, Molecular; D009068:Movement; D000072417:Protein Domains; D012326:RNA Splicing; D000072260:RNA Splicing Factors / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D020871:RNA Stability; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D016914:Ribonuclease H / Q000737:chemistry / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000201:enzymology; D017381:Spliceosomes / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*
chemical_list: C112200:CDC40 protein, human; D018797:Cell Cycle Proteins; C115374:PRPF8 protein, human; D000072260:RNA Splicing Factors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D017411:Ribonucleoproteins, Small Nuclear; D016914:Ribonuclease H; D053487:DEAD-box RNA Helicases; C545297:DHX8 protein, human; D000241:Adenosine