title: Crystal structure of a core spliceosomal protein interface.
abstract: The precise excision of introns from precursor mRNAs (pre-mRNAs) in eukaryotes is accomplished by the spliceosome, a complex assembly containing five small nuclear ribonucleoprotein (snRNP) particles. Human p14, a component of the spliceosomal U2 and U11/U12 snRNPs, has been shown to associate directly with the pre-mRNA branch adenosine early in spliceosome assembly and within the fully assembled spliceosome. Here we report the 2.5-A crystal structure of a complex containing p14 and a peptide derived from the p14-associated U2 snRNP component SF3b155. p14 contains an RNA recognition motif (RRM), the surface of which is largely occluded by a C-terminal alpha-helix and a portion of the SF3b155 peptide. An analysis of RNA.protein crosslinking to wild-type and mutant p14 shows that the branch adenosine directly interacts with a conserved aromatic within a pocket on the surface of the complex. This result, combined with a comparison of the structure with known RRMs and pseudoRRMs as well as model-building by using the electron cryomicroscopy structure of a spliceosomal U11/U12 di-snRNP, suggests that p14.SF3b155 presents a noncanonical surface for RNA recognition at the heart of the mammalian spliceosome.
authors.affiliation: Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2H7.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Schellenberg
authors.forename: Ross A
authors.initials: RA
authors.lastname: Edwards
authors.forename: Dustin B
authors.initials: DB
authors.lastname: Ritchie
authors.forename: Oliver A
authors.initials: OA
authors.lastname: Kent
authors.forename: Monika M
authors.initials: MM
authors.lastname: Golas
authors.forename: Holger
authors.initials: H
authors.lastname: Stark
authors.forename: Reinhard
authors.initials: R
authors.lastname: Lührmann
authors.forename: J N Mark
authors.initials: JN
authors.lastname: Glover
authors.forename: Andrew M
authors.initials: AM
authors.lastname: MacMillan
mesh_terms: D000241:Adenosine / Q000737:chemistry; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D003432:Cross-Linking Reagents / Q000494:pharmacology; D020285:Cryoelectron Microscopy; D018360:Crystallography, X-Ray; D006801:Humans; D007438:Introns; D008854:Microscopy, Electron; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry; D012313:RNA / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry; D017413:Ribonucleoprotein, U2 Small Nuclear / Q000737:chemistry; D017411:Ribonucleoproteins, Small Nuclear / Q000737:chemistry*; D017381:Spliceosomes / Q000737:chemistry* / Q000378:metabolism
chemical_list: D003432:Cross-Linking Reagents; D010455:Peptides; D011506:Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear; C098762:U11 small nuclear ribonucleoprotein, human; C552126:U11-U12 small nuclear ribonucleoprotein; C437317:U2 small nuclear ribonucleoprotein p14; D012313:RNA; D000241:Adenosine