High Resolution Structure of the Large Ribosomal Subunit from a Mesophilic Eubacterium

We describe the high resolution structure of the large ribosomal subunit from Deinococcus radiodurans (D50S), a gram-positive mesophile suitable for binding of antibiotics and functionally relevant ligands. The over-all structure of D50S is similar to that from the archae bacterium Haloarcula marismortui (H50S); however, a detailed comparison revealed significant differences, for example, in the orientation of nucleotides in peptidyl transferase center and in the structures of many ribosomal proteins. Analysis of ribosomal features involved in dynamic aspects of protein biosynthesis that are partially or fully disordered in H50S revealed the conformations of intersubunit bridges in unbound subunits, suggesting how they may change upon subunit association and how movements of the L1-stalk may facilitate the exit of tRNA.

[1]  J. Ballesta,et al.  Three‐dimensional cryo‐electron microscopy localization of EF2 in the Saccharomyces cerevisiae 80S ribosome at 17.5 Å resolution , 2000, The EMBO journal.

[2]  B. Golden,et al.  Ribosomal protein L6: structural evidence of gene duplication from a primitive RNA binding protein. , 1993, The EMBO journal.

[3]  Collaborative Computational,et al.  The CCP4 suite: programs for protein crystallography. , 1994, Acta crystallographica. Section D, Biological crystallography.

[4]  J. Frank,et al.  Localization of L11 protein on the ribosome and elucidation of its involvement in EF-G-dependent translocation. , 2001, Journal of molecular biology.

[5]  C. Vonrhein,et al.  Structure of the 30S ribosomal subunit , 2000, Nature.

[6]  H. Noller,et al.  Analysis of mutations at residues A2451 and G2447 of 23S rRNA in the peptidyltransferase active site of the 50S ribosomal subunit , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[7]  T. Steitz,et al.  The structural basis of ribosome activity in peptide bond synthesis. , 2000, Science.

[8]  Joachim Frank,et al.  Visualization of Trna Movements on the Escherichia coli 70s Ribosome during the Elongation Cycle , 2000, The Journal of cell biology.

[9]  D. Sabatini,et al.  CONTROLLED PROTEOLYSIS OF NASCENT POLYPEPTIDES IN RAT LIVER CELL FRACTIONS , 1970, The Journal of cell biology.

[10]  B. Hapke,et al.  Structural dynamics of bacterial ribosomes. I. Characterization of vacant couples and their relation to complexed ribosomes. , 1973, Journal of molecular biology.

[11]  Richard Brimacombe,et al.  The Database of Ribosomal Cross-links: an update , 1999, Nucleic Acids Res..

[12]  S. Tishchenko,et al.  Structure of ribosomal protein L30 from Thermus thermophilus at 1.9 A resolution: conformational flexibility of the molecule. , 1998, Acta crystallographica. Section D, Biological crystallography.

[13]  A. Liljas,et al.  The crystal structure of ribosomal protein L22 from Thermus thermophilus: insights into the mechanism of erythromycin resistance. , 1998, Structure.

[14]  V. Ramakrishnan,et al.  RIBOSOMAL PROTEIN L9 , 1997 .

[15]  S. Grzesiek,et al.  The RNA binding domain of ribosomal protein L11: three-dimensional structure of the RNA-bound form of the protein and its interaction with 23 S rRNA. , 1997, Journal of molecular biology.

[16]  G. Bricogne,et al.  [27] Maximum-likelihood heavy-atom parameter refinement for multiple isomorphous replacement and multiwavelength anomalous diffraction methods. , 1997, Methods in enzymology.

[17]  S. Douthwaite,et al.  Ribosomal proteins L11 and L10.(L12)4 and the antibiotic thiostrepton interact with overlapping regions of the 23 S rRNA backbone in the ribosomal GTPase centre. , 1993, Journal of molecular biology.

[18]  M. Wahl,et al.  Crystal structure of ribosomal protein L4 shows RNA‐binding sites for ribosome incorporation and feedback control of the S10 operon , 2000, The EMBO journal.

[19]  R. Garrett,et al.  Mapping important nucleotides in the peptidyl transferase centre of 23 S rRNA using a random mutagenesis approach. , 1995, Journal of molecular biology.

[20]  A Yonath,et al.  Structure of functionally activated small ribosomal subunit at 3.3 angstroms resolution. , 2000, Cell.

[21]  A. Liljas,et al.  Crystal structure of the RNA binding ribosomal protein L1 from Thermus thermophilus. , 1996, The EMBO journal.

[22]  T. Steitz,et al.  Structure of Escherichia coli ribosomal protein L25 complexed with a 5S rRNA fragment at 1.8-A resolution. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[23]  M van Heel,et al.  The 3D arrangement of the 23 S and 5 S rRNA in the Escherichia coli 50 S ribosomal subunit based on a cryo-electron microscopic reconstruction at 7.5 A resolution. , 2000, Journal of molecular biology.

[24]  Poul Nissen,et al.  Placement of protein and RNA structures into a 5 Å-resolution map of the 50S ribosomal subunit , 1999, Nature.

[25]  RIBOSOMAL PROTEIN L6 , 1999 .

[26]  T. Earnest,et al.  Crystal Structure of the Ribosome at 5.5 Å Resolution , 2001, Science.

[27]  Z. Otwinowski,et al.  [20] Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[28]  A. Rich,et al.  Partial resistance of nascent polypeptide chains to proteolytic digestion due to ribosomal shielding. , 1967, Journal of molecular biology.

[29]  V. Ramakrishnan,et al.  Structure of the 30 S ribosomal subunit , 2022 .

[30]  D. Draper,et al.  Recognition of the highly conserved GTPase center of 23 S ribosomal RNA by ribosomal protein L11 and the antibiotic thiostrepton. , 1991, Journal of molecular biology.

[31]  R. Garrett,et al.  Chloramphenicol resistance mutations in the single 23S rRNA gene of the archaeon Halobacterium halobium , 1991, Journal of bacteriology.

[32]  J. McCutcheon,et al.  A Detailed View of a Ribosomal Active Site The Structure of the L11–RNA Complex , 1999, Cell.

[33]  J. Walleczek,et al.  Comparative cross-linking study on the 50S ribosomal subunit from Escherichia coli. , 1989, Biochemistry.

[34]  F. Schluenzen,et al.  Structural basis for the interaction of antibiotics with the peptidyl transferase centre in eubacteria , 2001, Nature.

[35]  J. Wower,et al.  Ribosomal Protein L27 Participates in both 50 S Subunit Assembly and the Peptidyl Transferase Reaction* , 1998, The Journal of Biological Chemistry.

[36]  B. Cooperman,et al.  [3H]-p-azidopuromycin photoaffinity labeling of Escherichia coli ribosomes: evidence for site-specific interaction at U-2504 and G-2502 in domain V of 23S ribosomal RNA. , 1988, Biochemistry.

[37]  J Frank,et al.  Haloarcula marismortui 50S subunit-complementarity of electron microscopy and X-Ray crystallographic information. , 1999, Journal of structural biology.

[38]  T. Härd,et al.  The solution structure of ribosomal protein L36 from Thermus thermophilus reveals a zinc-ribbon-like fold. , 2000, Journal of molecular biology.

[39]  A Yonath,et al.  A tunnel in the large ribosomal subunit revealed by three-dimensional image reconstruction. , 1987, Science.

[40]  D. Draper,et al.  Contributions of basic residues to ribosomal protein L11 recognition of RNA. , 2000, Journal of molecular biology.

[41]  S. Segawa,et al.  End of the beginning , 1990, Nature.

[42]  S. Salzberg,et al.  Genome sequence of the radioresistant bacterium Deinococcus radiodurans R1. , 1999, Science.

[43]  J. Zou,et al.  Improved methods for building protein models in electron density maps and the location of errors in these models. , 1991, Acta crystallographica. Section A, Foundations of crystallography.

[44]  H. Urlaub,et al.  Peptide Environment of the Peptidyl Transferase Center from Escherichia coli 70 S Ribosomes as Determined by Thermoaffinity Labeling with Dihydrospiramycin (*) , 1995, The Journal of Biological Chemistry.

[45]  N. Sonenberg,et al.  Mapping of Escherichia coli ribosomal components involved in peptidyl transferase activity. , 1973, Proceedings of the National Academy of Sciences of the United States of America.

[46]  S. Kawamura,et al.  Ribosomal protein L5 has a highly twisted concave surface and flexible arms responsible for rRNA binding. , 2001, RNA.

[47]  R. Garrett,et al.  Assembly of proteins and 5 S rRNA to transcripts of the major structural domains of 23 S rRNA. , 1998, Journal of molecular biology.

[48]  E. Lattman,et al.  Crystal structure of a conserved ribosomal protein-RNA complex. , 1999, Science.

[49]  S. Dorner,et al.  Mechanism of ribosomal peptide bond formation. , 2001, Science.

[50]  V. Ramakrishnan,et al.  The crystal structure of ribosomal protein L14 reveals an important organizational component of the translational apparatus. , 1996, Structure.

[51]  H. Wittmann Architecture of prokaryotic ribosomes. , 1983, Annual review of biochemistry.

[52]  R. Huber,et al.  Flexibility, conformational diversity and two dimerization modes in complexes of ribosomal protein L12 , 2000, The EMBO journal.

[53]  F. Schluenzen,et al.  Structure of Functionally Activated Small Ribosomal Subunit , 2000 .

[54]  T. Steitz,et al.  The complete atomic structure of the large ribosomal subunit at 2.4 A resolution. , 2000, Science.

[55]  Sydney Brenner,et al.  The End of the Beginning , 2000, Science.

[56]  F. Schluenzen,et al.  Structure of Functionally Activated Small Ribosomal Subunit at 3.3 Å Resolution , 2000, Cell.

[57]  R. Milligan,et al.  Location of exit channel for nascent protein in 80S ribosome , 1986, Nature.

[58]  M. Stark,et al.  Ribosomes in thiostrepton-resistant mutants of Bacillus megaterium lacking a single 50 S subunit protein. , 1979, Journal of molecular biology.

[59]  A Yonath,et al.  Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3 , 2001, The EMBO journal.

[60]  M. Selmer,et al.  Structure of ribosomal protein TL5 complexed with RNA provides new insights into the CTC family of stress proteins. , 2001, Acta crystallographica. Section D, Biological crystallography.

[61]  A. Liljas,et al.  The end of the beginning: structural studies of ribosomal proteins. , 2000, Current opinion in structural biology.

[62]  J. Navaza,et al.  AMoRe: an automated package for molecular replacement , 1994 .

[63]  V. Ramakrishnan,et al.  Ribosomal protein L9: a structure determination by the combined use of X-ray crystallography and NMR spectroscopy. , 1996, Journal of molecular biology.

[64]  M. Lu,et al.  5S rRNA断片と複合体形成した大腸菌リボソーム蛋白質L25の1.8Å分解能での構造 , 2000 .

[65]  Harry F. Noller,et al.  Interaction of antibiotics with functional sites in 16S ribosomal RNA , 1987, Nature.

[66]  E. Dabbs,et al.  Functional studies on ribosomes lacking protein L1 from mutant Escherichia coli. , 1980, European journal of biochemistry.

[67]  Hilla Peretz,et al.  Ju n 20 03 Schrödinger ’ s Cat : The rules of engagement , 2003 .

[68]  S. Dorner,et al.  A conformational change in the ribosomal peptidyl transferase center upon active/inactive transition , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[69]  J. Abrahams,et al.  Methods used in the structure determination of bovine mitochondrial F1 ATPase. , 1996, Acta crystallographica. Section D, Biological crystallography.

[70]  A. Mankin,et al.  Ribosomal peptidyl transferase can withstand mutations at the putative catalytic nucleotide , 2001, Nature.