title: Ribosomal protein L9: a structure determination by the combined use of X-ray crystallography and NMR spectroscopy.
abstract: The structure of protein L9 from the Bacillus stearothernophilus ribosome has been determined at 2.5 A resolution by refinement against single crystal X-ray diffraction data with additional constraints provided by NMR data. This highly elongated protein consists of two domains separated by a nine-turn connecting helix. Conserved aromatic and positively charged amino acid residues on the surface of each domain are likely to be directly involved in binding 23 S ribosomal RNA. The shape of the protein, with its two widely spaced RNA-binding sites, suggests that it may serve as a "molecular strut", most likely playing a role in ribosome assembly and/or maintaining the catalytically active conformation of the ribosomal RNA. The combined use of X-ray and NMR data in the refinement procedure was essential in defining the N-terminal domain of the protein, which was relatively poorly determined by the X-ray data alone. In addition to resolving the ambiguities in defining the hydrophobic core and side-chain conformations with the N-terminal domain, this combined NMR-X-ray analysis provides the first detailed and accurate view of the N-terminal RNA-binding site. NMR data also showed that the N-terminal domain is stable in solution, indicated by amide protons that are protected from solvent exchange. The lack of definition of the N-terminal domain in the X-ray structure is therefore likely due to packing disorder within the crystal rather than structural instability. This combined NMR-X-ray analysis provides a useful model as to how X-ray and NMR data can be practically and logically combined in the determination of the structure of a single protein molecule.
authors.affiliation: Department of Chemistry and Biochemistry, University of Texas at Austin, 78712, USA.
authors.forename: D W
authors.initials: DW
authors.lastname: Hoffman
authors.forename: C S
authors.initials: CS
authors.lastname: Cameron
authors.forename: C
authors.initials: C
authors.lastname: Davies
authors.forename: S W
authors.initials: SW
authors.lastname: White
authors.forename: V
authors.initials: V
authors.lastname: Ramakrishnan
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D018360:Crystallography, X-Ray / Q000379:methods; D001411:Geobacillus stearothermophilus / Q000737:chemistry / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D011487:Protein Conformation; D012329:RNA, Bacterial / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D001426:Bacterial Proteins; D012329:RNA, Bacterial; D011994:Recombinant Proteins; D012269:Ribosomal Proteins; C049650:ribosomal protein L9