title: A portable RNA sequence whose recognition by a synthetic antibody facilitates structural determination.
abstract: RNA crystallization and phasing represent major bottlenecks in RNA structure determination. Seeking to exploit antibody fragments as RNA crystallization chaperones, we have used an arginine-enriched synthetic Fab library displayed on phage to obtain Fabs against the class I ligase ribozyme. We solved the structure of a Fab-ligase complex at 3.1-Å resolution using molecular replacement with Fab coordinates, confirming the ribozyme architecture and revealing the chaperone's role in RNA recognition and crystal contacts. The epitope resides in the GAAACAC sequence that caps the P5 helix, and this sequence retains high-affinity Fab binding within the context of other structured RNAs. This portable epitope provides a new RNA crystallization chaperone system that easily can be screened in parallel to the U1A RNA-binding protein, with the advantages of a smaller loop and Fabs' high molecular weight, large surface area and phasing power.
authors.affiliation: Department of Chemistry, University of Chicago, Chicago, Illinois, USA.
authors.forename: Yelena
authors.initials: Y
authors.lastname: Koldobskaya
authors.forename: Erica M
authors.initials: EM
authors.lastname: Duguid
authors.forename: David M
authors.initials: DM
authors.lastname: Shechner
authors.forename: Nikolai B
authors.initials: NB
authors.lastname: Suslov
authors.forename: Jingdong
authors.initials: J
authors.lastname: Ye
authors.forename: Sachdev S
authors.initials: SS
authors.lastname: Sidhu
authors.forename: David P
authors.initials: DP
authors.lastname: Bartel
authors.forename: Shohei
authors.initials: S
authors.lastname: Koide
authors.forename: Anthony A
authors.initials: AA
authors.lastname: Kossiakoff
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Piccirilli
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D003460:Crystallization / Q000379:methods*; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D008025:Ligases / Q000737:chemistry / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D019151:Peptide Library; D016337:RNA, Catalytic / Q000737:chemistry* / Q000276:immunology; D022661:RNA, Untranslated / Q000737:chemistry
chemical_list: D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library; D016337:RNA, Catalytic; D022661:RNA, Untranslated; D008025:Ligases