title: The box C/D sRNP dimeric architecture is conserved across domain Archaea.
abstract: Box C/D small (nucleolar) ribonucleoproteins [s(no)RNPs] catalyze RNA-guided 2'-O-ribose methylation in two of the three domains of life. Recent structural studies have led to a controversy over whether box C/D sRNPs functionally assemble as monomeric or dimeric macromolecules. The archaeal box C/D sRNP from Methanococcus jannaschii (Mj) has been shown by glycerol gradient sedimentation, gel filtration chromatography, native gel analysis, and single-particle electron microscopy (EM) to adopt a di-sRNP architecture, containing four copies of each box C/D core protein and two copies of the Mj sR8 sRNA. Subsequently, investigators used a two-stranded artificial guide sRNA, CD45, to assemble a box C/D sRNP from Sulfolobus solfataricus with a short RNA methylation substrate, yielding a crystal structure of a mono-sRNP. To more closely examine box C/D sRNP architecture, we investigate the role of the omnipresent sRNA loop as a structural determinant of sRNP assembly. We show through sRNA mutagenesis, native gel electrophoresis, and single-particle EM that a di-sRNP is the near exclusive architecture obtained when reconstituting box C/D sRNPs with natural or artificial sRNAs containing an internal loop. Our results span three distantly related archaeal species--Sulfolobus solfataricus, Pyrococcus abyssi, and Archaeoglobus fulgidus--indicating that the di-sRNP architecture is broadly conserved across the entire archaeal domain.
authors: Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
authors: Kathleen R
authors: KR
authors: Bower-Phipps
authors: David W
authors: DW
authors: Taylor
authors: Hong-Wei
authors: HW
authors: Wang
authors: Susan J
authors: SJ
authors: Baserga
mesh_terms: D001105:Archaea / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D008745:Methylation; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D019642:RNA, Archaeal / Q000235:genetics / Q000378:metabolism*; D020537:RNA, Small Nucleolar / Q000235:genetics / Q000378:metabolism*; D020718:Ribonucleoproteins, Small Nucleolar / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D019843:Archaeal Proteins; D019642:RNA, Archaeal; D020537:RNA, Small Nucleolar; D020718:Ribonucleoproteins, Small Nucleolar