title: A conformational switch in PRP8 mediates metal ion coordination that promotes pre-mRNA exon ligation.
abstract: Splicing of pre-mRNAs in eukaryotes is catalyzed by the spliceosome, a large RNA-protein metalloenzyme. The catalytic center of the spliceosome involves a structure comprising the U2 and U6 snRNAs and includes a metal bound by U6 snRNA. The precise architecture of the splicesome active site, however, and the question of whether it includes protein components, remains unresolved. A wealth of evidence places the protein PRP8 at the heart of the spliceosome through assembly and catalysis. Here we provide evidence that the RNase H domain of PRP8 undergoes a conformational switch between the two steps of splicing, rationalizing yeast prp8 alleles that promote either the first or second step. We also show that this switch unmasks a metal-binding site involved in the second step. Together, these data establish that PRP8 is a metalloprotein that promotes exon ligation within the spliceosome.
authors.affiliation: Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Schellenberg
authors.forename: Tao
authors.initials: T
authors.lastname: Wu
authors.forename: Dustin B
authors.initials: DB
authors.lastname: Ritchie
authors.forename: Sebastian
authors.initials: S
authors.lastname: Fica
authors.forename: Jonathan P
authors.initials: JP
authors.lastname: Staley
authors.forename: Karim A
authors.initials: KA
authors.lastname: Atta
authors.forename: Paul
authors.initials: P
authors.lastname: LaPointe
authors.forename: Andrew M
authors.initials: AM
authors.lastname: MacMillan
mesh_terms: D018360:Crystallography, X-Ray; D007477:Ions / Q000378:metabolism*; D008026:Ligation; D008670:Metals / Q000378:metabolism*; D008958:Models, Molecular; D011487:Protein Conformation; D012322:RNA Precursors / Q000378:metabolism*; D016914:Ribonuclease H / Q000737:chemistry / Q000378:metabolism; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000737:chemistry* / Q000378:metabolism*; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000737:chemistry* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D007477:Ions; D008670:Metals; C077018:PRP8 protein, S cerevisiae; D012322:RNA Precursors; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; D016914:Ribonuclease H