title: Functional requirement for symmetric assembly of archaeal box C/D small ribonucleoprotein particles.
abstract: Box C/D small ribonucleoprotein particles (sRNPs) are archaeal homologs of small nucleolar ribonucleoprotein particles (snoRNPs) in eukaryotes that are responsible for site specific 2'-O-methylation of ribosomal and transfer RNAs. The function of box C/D sRNPs is characterized by step-wise assembly of three core proteins around a box C/D RNA that include fibrillarin, Nop5p, and L7Ae. The most distinct structural feature in all box C/D RNAs is the presence of two conserved box C/D motifs accompanied by often a single, and sometimes two, antisense elements located immediately upstream of either the D or D' box. Despite this asymmetric distribution of antisense elements, the bipartite feature of the box C/D motifs appears to be in pleasing agreement with a recently reported three-dimensional structure of the core protein complex between fibrillarin and Nop5p. This investigates functional implications of the symmetric features both in box C/D RNAs and in the fibrillarin-Nop5p complex. Site-directed mutagenesis was employed to generate box C/D RNAs lacking one of the two box C/D motifs and a mutant fibrillarin-Nop5p complex deficient in self-association. The ability of the mutated components to assemble and to direct methyl transfer reactions was assessed by gel mobility-shift, analytical ultracentrifugation, and in vitro catalysis studies. The results presented here suggest that, while a box C/D sRNP is capable of asymmetrical assembly, the symmetries in both the box C/D RNA and in the fibrillarin-Nop5p complex are required for efficient catalysis. These findings underscore the importance of functional assembly in methyl transfer reactions.
authors.affiliation: Department of Chemistry and Biochemistry, Institute of Molecular Biophysics, Florida State University, Tallahassee, FL 32306, USA.
authors.forename: Rumana
authors.initials: R
authors.lastname: Rashid
authors.forename: Mohamed
authors.initials: M
authors.lastname: Aittaleb
authors.forename: Qiong
authors.initials: Q
authors.lastname: Chen
authors.forename: Katharina
authors.initials: K
authors.lastname: Spiegel
authors.forename: Borries
authors.initials: B
authors.lastname: Demeler
authors.forename: Hong
authors.initials: H
authors.lastname: Li
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019611:Archaeoglobus fulgidus / Q000235:genetics / Q000378:metabolism*; D020029:Base Pairing; D001483:Base Sequence; D001665:Binding Sites; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D024202:Electrophoretic Mobility Shift Assay; D008745:Methylation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins; D017393:RNA Editing; D019642:RNA, Archaeal / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020537:RNA, Small Nucleolar / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020718:Ribonucleoproteins, Small Nucleolar / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D058727:RNA, Small Untranslated
chemical_list: D019843:Archaeal Proteins; D002868:Chromosomal Proteins, Non-Histone; D009687:Nuclear Proteins; D019642:RNA, Archaeal; D020537:RNA, Small Nucleolar; D020718:Ribonucleoproteins, Small Nucleolar; C047458:fibrillarin; D058727:RNA, Small Untranslated