A new model for the three-dimensional folding of Escherichia coli 16 S ribosomal RNA. II. The RNA-protein interaction data.

The map of the mass centres of the 21 proteins from the Escherichia coli 30 S ribosomal subunit, as determined by neutron scattering, was fitted to a cryoelectron microscopic (cryo-EM) model at a resolution of 20 A of 70 S ribosomes in the pre-translocational state, carrying tRNA molecules at the A and P sites. The fit to the 30 S moiety of the 70 S particles was accomplished with the help of the well-known distribution of the ribosomal proteins in the head, body and side lobe regions of the 30 S subunit, as determined by immuno electron microscopy (IEM). Most of the protein mass centres were found to lie close to the surface (or even outside) of the cryo-EM contour of the 30 S subunit, supporting the idea that the ribosomal proteins are arranged peripherally around the rRNA. The ribosomal protein distribution was then compared with the corresponding model for the 16 S rRNA, fitted to the same EM contour (described in an accompanying paper), in order to analyse the mutual compatibility of the arrangement of proteins and rRNA in terms of the available RNA-protein interaction data. The information taken into account included the hydroxyl radical and base foot-printing data from Noller's laboratory, and our own in situ cross-linking results. Proteins S1 and S14 were not considered, due to the lack of RNA-protein data. Among the 19 proteins analysed, 12 (namely S2, S4, S5, S7, S8, S9, S10, S11, S12, S15, S17 and S21) showed a fit to the rRNA model that varied from being excellent to at least acceptable. Of the remaining 7, S3 and S13 showed a rather poor fit, as did S18 (which is considered in combination with S6 in the foot-printing experiments). S16 was difficult to evaluate, as the foot-print data for this protein cover a large area of the rRNA. S19 and S20 showed a bad fit in terms of the neutron map, but their foot-print and cross-link sites were clustered into compact groups in the rRNA model in those regions of the 30 S subunit where these proteins have respectively been located by IEM studies.

[1]  V. Ramakrishnan,et al.  The structure of ribosomal protein S5 reveals sites of interaction with 16S rRNA , 1992, Nature.

[2]  R. Brimacombe,et al.  A detailed model of the three-dimensional structure of Escherichia coli 16 S ribosomal RNA in situ in the 30 S subunit. , 1988, Journal of molecular biology.

[3]  G. Kramer,et al.  Structure, Function, and Genetics of Ribosomes , 1986, Springer Series in Molecular Biology.

[4]  R. Crichton,et al.  Determination of the distribution of protein and nucleic acid in the 70 S ribosomes of Escherichia coli and their 30 S subunits by neutron scattering. , 1978, Journal of molecular biology.

[5]  R. Brimacombe,et al.  A new model for the three-dimensional folding of Escherichia coli 16 S ribosomal RNA. I. Fitting the RNA to a 3D electron microscopic map at 20 A. , 1997, Journal of molecular biology.

[6]  V. Ramakrishnan,et al.  Distribution of protein and RNA in the 30S ribosomal subunit. , 1986, Science.

[7]  J. Ofengand,et al.  Structural and Functional Interactions of the tRNA-Ribosome Complex , 1986 .

[8]  J. Lake,et al.  Ribosome Structure, Function, and Evolution: Mapping Ribosomal RNA, Proteins, and Functional Sites in Three Dimensions , 1986 .

[9]  R B Altman,et al.  Computational methods for defining the allowed conformational space of 16S rRNA based on chemical footprinting data. , 1996, RNA.

[10]  R. Albrecht-Ehrlich,et al.  Immunoelectron microscopic localization of ribosomal proteins BS8, BS9, BS20, BL3 and BL21 on the surface of 30S and 50S subunits from Bacillus stearothermophilus. , 1993, European journal of biochemistry.

[11]  R. Brimacombe,et al.  Getting closer to an understanding of the three-dimensional structure of ribosomal RNA. , 1995, Biochemistry and cell biology = Biochimie et biologie cellulaire.

[12]  Helena Berglund,et al.  Solution structure of the ribosomal RNA binding protein S15 from Thermus thermophilus , 1997, Nature Structural Biology.

[13]  S C Harvey,et al.  A quantitative model of the Escherichia coli 16 S RNA in the 30 S ribosomal subunit. , 1994, Journal of molecular biology.

[14]  J. Ofengand,et al.  Covalent crosslinking of tRNA1Val to 16S RNA at the ribosomal P site: identification of crosslinked residues. , 1982, Proceedings of the National Academy of Sciences of the United States of America.

[15]  R. Batey,et al.  Interaction of the Bacillus stearothermophilus ribosomal protein S15 with 16 S rRNA: II. Specificity determinants of RNA-protein recognition. , 1996, Journal of molecular biology.

[16]  R. Brimacombe.  RNA-protein interactions in the Escherichia coli ribosome. , 1991, Biochimie.

[17]  M Kjeldgaard,et al.  Positions of S2, S13, S16, S17, S19 and S21 in the 30 S ribosomal subunit of Escherichia coli. , 1988, Journal of molecular biology.

[18]  I. Tanaka,et al.  Crystal structure of a prokaryotic ribosomal protein. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[19]  C. Gualerzi,et al.  Identification of the amino acid residues of proteins S5 and S8 adjacent to each other in the 30 S ribosomal subunit of Escherichia coli. , 1979, The Journal of biological chemistry.

[20]  E. Westhof,et al.  Role of conserved nucleotides in building the 16S rRNA binding site of E. coli ribosomal protein S8. , 1994, Nucleic acids research.

[21]  B. Cooperman,et al.  Ribosomal components neighboring the conserved 518-533 loop of 16S rRNA in 30S subunits. , 1994, Biochemistry.

[22]  J. Frank,et al.  Three-dimensional reconstruction of the 70S Escherichia coli ribosome in ice: the distribution of ribosomal RNA , 1991, The Journal of cell biology.

[23]  R. Brimacombe,et al.  The ribosomal environment of tRNA: crosslinks to rRNA from positions 8 and 20:1 in the central fold of tRNA located at the A, P, or E site. , 1995, RNA: A publication of the RNA Society.

[24]  V. Ramakrishnan,et al.  Structural evidence for specific S8-RNA and S8-protein interactions within the 30S ribosomal subunit: ribosomal protein S8 from Bacillus stearothermophilus at 1.9 A resolution. , 1996, Structure.

[25]  R. Brimacombe,et al.  The localization of multiple sites on 16S RNA which are cross-linked to proteins S7 and S8 in Escherichia coli 30S ribosomal subunits by treatment with 2-iminothiolane. , 1983, Nucleic acids research.

[26]  B. Cooperman,et al.  Immune electron microscopic localization of dinitrophenyl-modified ribosomal protein S19 in reconstituted Escherichia coli 30 S subunits using antibodies to dinitrophenol. , 1988, Journal of Biological Chemistry.

[27]  R. Brimacombe,et al.  The structure of ribosomal RNA: a three-dimensional jigsaw puzzle. , 1995, European journal of biochemistry.

[28]  Ribosomal protein S17: characterization of the three-dimensional structure by 1H and 15N NMR. , 1993 .

[29]  J. Wower,et al.  Probing tRNA binding sites on the Escherichia coli 30 S ribosomal subunit with photoreactive analogs of the anticodon arm. , 1990, Biochimica et biophysica acta.

[30]  B. Wittmann-Liebold.  Ribosomal Proteins: Their Structure and Evolution , 1986 .

[31]  H. Noller,et al.  Hydroxyl radical footprinting of ribosomal proteins on 16S rRNA. , 1995, RNA.

[32]  R. Brimacombe,et al.  Identification and Sequence Analysis of Contact Sites between Ribosomal Proteins and rRNA in Escherichia coli 30 S Subunits by a New Approach Using Matrix-assisted Laser Desorption/Ionization-Mass Spectrometry Combined with N-terminal Microsequencing* , 1997, The Journal of Biological Chemistry.

[33]  R. Brimacombe,et al.  The ribosomal neighbourhood of the central fold of tRNA: cross-links from position 47 of tRNA located at the A, P or E site. , 1995, Nucleic acids research.

[34]  Walter E. Hill,et al.  The Ribosome : structure, function, and evolution , 1990 .

[35]  Protein binding sites on Escherichia coli 16S ribosomal RNA; RNA regions that are protected by proteins S7, S9 and S19, and by proteins S8, S15 and S17. , 1988, Nucleic acids research.

[36]  H. Noller.  Structure of ribosomal RNA. , 1984, Annual review of biochemistry.

[37]  H. Noller,et al.  Site-Directed Hydroxyl Radical Probing of the rRNA Neighborhood of Ribosomal Protein S5 , 1996, Science.

[38]  L. Brakier-Gingras,et al.  Interaction of Escherichia coli ribosomal protein S7 with 16S rRNA. , 1993, Nucleic acids research.

[39]  V. Ramakrishnan,et al.  Crystal structure of prokaryotic ribosomal protein L9: a bi‐lobed RNA‐binding protein. , 1994, The EMBO journal.

[40]  M van Heel,et al.  A new model for the three-dimensional folding of Escherichia coli 16 S ribosomal RNA. III. The topography of the functional centre. , 1997, Journal of molecular biology.

[41]  R. Brimacombe,et al.  RNA-protein cross-linking in Escherichia coli ribosomal subunits: localization of sites on 16S RNA which are cross-linked to proteins S17 and S21 by treatment with 2-iminothiolane. , 1986, Nucleic acids research.

[42]  R. Brimacombe,et al.  RNA-protein cross-linking in Escherichia coli 30S ribosomal subunits; determination of sites on 16S RNA that are cross-linked to proteins S3, S4, S7, S9, S10, S11, S17, S18 and S21 by treatment with bis-(2-chloroethyl)-methylamine. , 1987, Nucleic acids research.

[43]  R. Brimacombe,et al.  RNA-protein cross-linking in Escherichia coli 30S ribosomal subunits; determination of sites on 16S RNA that are cross-linked to proteins S3, S4, S5, S7, S8, S9, S11, S13, S19 and S21 by treatment with methyl p-azidophenyl acetimidate. , 1987, Nucleic acids research.

[44]  R. Brimacombe,et al.  Arrangement of tRNAs in Pre- and Posttranslocational Ribosomes Revealed by Electron Cryomicroscopy , 1997, Cell.

[45]  H. Noller,et al.  Selective perturbation of G530 of 16 S rRNA by translational miscoding agents and a streptomycin-dependence mutation in protein S12. , 1994, Journal of molecular biology.

[46]  J. Wower,et al.  Mapping the central fold of tRNA2(fMet) in the P site of the Escherichia coli ribosome. , 1993, Biochemistry.

[47]  C. Zwieb,et al.  RNA-protein cross-linking in Escherichia coli 30s ribosomal subunits: precise localisation of the nucleotide in 16s RNA which is coupled to protein S7 by ultraviolet irradiation , 1979 .

[48]  H. Noller,et al.  Model for the three-dimensional folding of 16 S ribosomal RNA. , 1988, Journal of molecular biology.