Mapping the three-dimensional locations of ribosomal RNA and proteins

Scheinman, A.; Atha, T.; Aguinaldo, Anna-Marie; Kahan, Lawrence; Shankweiler, G.; Lake, James A.

doi:10.1016/0300-9084(92)90108-q

Cited by 23 publications

(10 citation statements)

References 40 publications

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“…We found the location by comparing the 30S subunit portion of that map, which contains S1, with the x-ray map of the S1-depleted T. thermophilus 30S subunit (25). The position derived from this study is consistent with results from IEM (17,18,19), neutron scattering (11,20,21), and S1-rRNA crosslinking (22) experiments, and it helps to explain all existing biochemical data (27,28). Our study shows that S1 is an elongated structure that makes several contacts with components of head, platform, and main body of the 30S subunit.…”

supporting

confidence: 79%

Visualization of protein S1 within the 30S ribosomal subunit and its interaction with messenger RNA

Sengupta

Agrawal

Frank

2001

Proc. Natl. Acad. Sci. U.S.A.

148

115

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S1 is the largest ribosomal protein, present in the small subunit of the bacterial ribosome. It has a pivotal role in stabilizing the mRNA on the ribosome. Thus far, S1 has eluded structural determination. We have identified the S1 protein mass in the cryo-electron microscopic map of the Escherichia coli ribosome by comparing the map with a recent x-ray crystallographic structure of the 30S subunit, which lacks S1. According to our finding, S1 is located at the junction of head, platform, and main body of the 30S subunit, thus explaining all existing biochemical and crosslinking data. Protein S1 as identified in our map has a complex, elongated shape with two holes in its central portion. The N-terminal domain, forming one of the extensions, penetrates into the head of the 30S subunit. Evidence for direct interaction of S1 with 11 nucleotides of the mRNA, immediately upstream of the Shine-Dalgarno sequence, explains the protein's role in the recognition of the 5 region of mRNA. P rotein S1 is the largest ribosomal protein, 68 kDa (1), present in the small subunit of the Escherichia coli 70S ribosome. It has been implicated in the selection of the translation start site on the 30S subunit (2). It has been suggested that a pyrimidinerich region upstream of the Shine-Dalgarno (SD) sequence of mRNA interacts with protein S1 and serves as one of the ribosome recognition sites (3). Protein S1 has been reported to be necessary in some cases for translation initiation (4) and for translation elongation (5). It is the only ribosomal protein that has a high affinity for mRNA (6). As a ribosomal protein, S1 is strikingly atypical. Association of S1 to the ribosome is weak and reversible (ribosome preparations often contain less than stoichiometric amounts of protein S1; ref. 7), whereas most other ribosomal proteins are strongly bound. Furthermore, S1 is an acidic protein, whereas all other ribosomal proteins, with the exception of protein L7͞L12, are basic.The S1 protein consists of 557 amino acid residues. Mild trypsin digestion cleaves S1 into two fragments (8), whereas cleavage with cyanogen bromide yields three different fragments (9). The N-terminal fragment from the latter cleavage contains amino acids 1-193, the middle fragment contains amino acids 224-309, and the C-terminal fragment contains amino acids 332-547 (9). The N-terminal domain binds to the platform region of the 30S subunit (10), whereas the C-terminal domain has been suggested to interact with RNAs (11). S1 has been reported to have an elongated shape in solution (12, 13). Neutron scattering (11) and fluorescence anisotropy experiments (14) indicate that the protein maintains its elongated shape even in its ribosome-bound state.So far, no x-ray crystallographic study has been reported for S1. However, an NMR structure of a 75-aa fragment of S1-type polynucleotide phosphorylase of E. coli, which corresponds to the C-terminal domain of S1, has been determined (15). The structure consists of a five-stranded antiparallel ␤-barrel, a structural motif known...

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supporting

confidence: 79%

Visualization of protein S1 within the 30S ribosomal subunit and its interaction with messenger RNA

Sengupta

Agrawal

Frank

2001

Proc. Natl. Acad. Sci. U.S.A.

148

115

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“…9C). Scheinman et al (1992) have similarly noted a site for S20 near to the base of the subunit. Even allowing for the fact that the neutron-scattering data define the position of the mass centre of a protein, whereas IEM identifies sites on the protein surface, this separation is clearly too large for a small protein such as S20.…”

Section: Discussionmentioning

confidence: 84%

“…Traut et al, 1980). Scheinman et al (1992) have reported a localization of E. coEi S9, which also lay in thc head of the 30s subunit, close to or overlapping the site for S7. However, their S9 site was on the interface side of the subunit.…”

Section: Discussionmentioning

confidence: 86%

Immunoelectron microscopic localization of ribosomal proteins BS8, BS9, BS20, BL3 and BL21 on the surface of 30S and 50S subunits from Bacillus stearothermophilus

Schwedler

Albrecht-Ehrlich

Rak

1993

European Journal of Biochemistry

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The locations of ribosomal proteins BS8, BS9 and BS20 on the 30s subunit of Bacillus stearothermophilus ribosomes, and of BL3 and BL21 on the 50s subunit, were determined by immunoelectron microscopy. BL3 was found to lie half-way down the body of the 50s subunit on the interface side, below the L7L12 stalk, in agreement with the placement of the corresponding protein in Escherichia coli by neutron-scattering; BL21 was located at a similar position on the solvent side of the subunit, as predicted by cross-linking experiments with E. coli ribosomes. Similarly, BS8 was found in the upper region of the body of the 30s subunit on the solvent side, and BS9 on the top of the head of the subunit, also on the solvent side, both positions being in good agreement with neutron-scattering data and other immunoelcctron microscopy results. In contrast, BS20 was found to lie at the extreme base of the body of the 30s subunit; this placement is not compatible with the location of E. coli S20 by neutron-scattering but fits very plausibly with other biochemical data, such as sites of RNA-protein footprinting on 16s RNA, relating to the location of S20 inImmunoelectron microscopy (IEM) is a long-established method for investigating the distribution of individual ribosomal proteins on the surface of the ribosomal subunits. In the case of the 30s subunit from Eschericlzia coli, our laboratory (Stoffler-Meilicke and Stoffler, 1990) and others (Oakes et al., 1990) have been able to locate epitopes for the majority of the small subunit ribosomal proteins with the help of this technique; the results so far have shown a high level of agreement with the neutron-scattering results of Capel et al. (1988), who have mapped the positions of the mass centers of all 21 of the 30s subunit proteins. In the case of the 50s subunit, the data sets are far less complete, although epitopes for more than half of the proteins have been located by IEM (Stoffler-Meilicke and Stoffler, 1990) and the mass centers of seven proteins have been mapped by neutron-scattering (May et al., 1992). A preliminary model for the positions of 29 of the 50s subunit proteins has been proposed (Walleczek et al., 1988), by combining the IEM data with the results of in siru protcin-protein cross-linking experiments.The more recent IEM studies have turned to the investigation of ribosomal proteins from Bacillus srearothermophi-/us. The overall morphology of ribosomal subunits from this organism is very similar to that observed in E. coli (e.g. Van Heel and Stoffler-Meilicke, 1985). Furthermore, the ribosomal proteins show a high degree of sequence similarity to their counterparts in E. coli (Wittmann-Liebold, 1988) cases so far examined it has been demonstrated that the IEM locations of similar proteins on the ribosomal subunit surfaces are identical in the two organisms (Stoffler-Meilicke et al., 1984;Stoffler and Stoffler-Meilicke, 1986; Hack1 and Stoffler-Meilicke, 1988). This finding has two important consequences. First, it shows that the sequence similarity is indeed r...

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“…Thus antibodies can be used as protein-specific stains to determine the positions of protein epitopes on the surface of the ribosome. By the mid 1980s, the positions of the surface epitopes of all the proteins in the E. coli small subunit had been mapped this way, as well as those of many large-subunit proteins (70,77). 5S rRNA, some critical rRNA sequences, and a number of other important sites on the ribosome had also been located (e.g.…”

Section: Quaternary Structurementioning

confidence: 99%

The Three-Dimensional Structure of the Ribosome and Its Components

Moore

1998

Annu. Rev. Biophys. Biomol. Struct.

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Exciting progress has been made in the last decade by those who use physical methods to study the structure of the ribosome and its components. The structures of 10 ribosomal proteins and three isolated ribosomal protein domains are known, and the conformations of a significant number of rRNA sequences have been determined. Electron microscopists have made major advances in the analysis of images of ribosomes, and microscopically derived ribosome models at resolutions approaching 10A are likely quite soon. Furthermore, ribosome crystallographers are on the verge of phasing the diffraction patterns they have had for several years, and near-atomic resolution models for entire ribosomal subunits could emerge from this source at any time. The literature relevant to these developments is reviewed below.

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Mapping the three-dimensional locations of ribosomal RNA and proteins

Cited by 23 publications

References 40 publications

Visualization of protein S1 within the 30S ribosomal subunit and its interaction with messenger RNA

Visualization of protein S1 within the 30S ribosomal subunit and its interaction with messenger RNA

Immunoelectron microscopic localization of ribosomal proteins BS8, BS9, BS20, BL3 and BL21 on the surface of 30S and 50S subunits from Bacillus stearothermophilus

The Three-Dimensional Structure of the Ribosome and Its Components

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