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Chapter 1 : Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit

Authors: V. Ramakrishnan1, Malcolm S. Capel3, William M. Clemons, Jr.4, Joanna L. C. May4, Brian T. Wimberly4
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Affiliations: 1: Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84132; 2: Structural Studies Division, MRC Laboratory of Molecular Biology, Hills Road, CambridgeCB2 2QH, England; 3: Biology Department, Brookhaven National Laboratory, Upton, NY 11973.; 4: Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84132
Content Type: Monograph
Publication Year: 2000

Category: Microbial Genetics and Molecular Biology

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Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, Page 1 of 2

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Abstract:

In 1991, Yonath and coworkers, after nearly a decade of pioneering work on the crystallization of ribosomes, showed that it was possible to obtain diffraction to beyond 3 Å from crystals of the 50S subunit of Haloarcula marismortui. This work marked a milestone in ribosome crystallography, because it established that in principle an atomic-resolution structure of a ribosomal subunit could be obtained. In the meantime, work on whole-ribosome crystallography was also carried out with . The problem of nonisomorphism can be alleviated by the use of multiwavelength anomalous dispersion, in which phase information is obtained by data collection on the same crystal at different wavelengths. Alpha helices of proteins are also visible in the structure, so that it is possible to find and to determine the orientations of proteins of known crystal structure in the map. It is possible to see protein-RNA complexes in many cases. It was possible to determine a fold for it even though the resolution is well below the traditional “atomic” resolution of 3.5 Å required for a new trace because the central domain contains a high density of proteins of previously known crystal structure and well-characterized biochemistry. It is tempting to model much of the ribosome at an intermediate resolution, where one can see double-stranded RNA and recognize known protein structures, or at even-lower resolution, as has been done by others in conjunction with biochemical and electron microscopic data.

Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1

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Double-Stranded RNA
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Haloarcula marismortui
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Thermus thermophilus
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Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit
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Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1
/content/10.1128/9781555818142.chap1.fig1-1
Figure 1

Harker section ( = 0.5) of an isomorphous difference Patterson map from a W17 derivative of 30S crystals, showing both a major and a minor site.

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Image of Figure 1
Figure 1

Harker section ( = 0.5) of an isomorphous difference Patterson map from a W17 derivative of 30S crystals, showing both a major and a minor site.

Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1
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Figure 2

Electron density map at 5.5-Å resolution. (a) A stretch of double-stranded RNA that shows individual strands, major and minor grooves, and bumps that correspond to phosphate groups. (b) Fit of the crystal structure of S6 ( ) to the electron density, showing that individual proteins of known structure can be both located and oriented in the map.

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10.1128/9781555818142/fig1-2.gif
Image of Figure 2
Figure 2

Electron density map at 5.5-Å resolution. (a) A stretch of double-stranded RNA that shows individual strands, major and minor grooves, and bumps that correspond to phosphate groups. (b) Fit of the crystal structure of S6 ( ) to the electron density, showing that individual proteins of known structure can be both located and oriented in the map.

Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1
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Figure 3

(a) Stereo view of the fold of the central domain of 16S RNA. (b) Stereo view of the three-helix junction formed by helices 20, 21, and 22 in the central domain, with associated proteins. The green helix shows the interaction of the N-terminal helix of S8 with a minor groove of H25.

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10.1128/9781555818142/fig1-3.gif
Image of Figure 3
Figure 3

(a) Stereo view of the fold of the central domain of 16S RNA. (b) Stereo view of the three-helix junction formed by helices 20, 21, and 22 in the central domain, with associated proteins. The green helix shows the interaction of the N-terminal helix of S8 with a minor groove of H25.

Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1
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Figure 4

Relative positions of RNA helices 24, 27, and 44 (the 1400/1500 stem-loop) in the structure.

10.1128/9781555818142/fig1-4_thmb.gif
10.1128/9781555818142/fig1-4.gif
Image of Figure 4
Figure 4

Relative positions of RNA helices 24, 27, and 44 (the 1400/1500 stem-loop) in the structure.

Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1
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Figure 5

Importance of protein-protein interactions in the 30S subunit. The figure shows relative positions of proteins S4, S5, and S8 in the 30S structure. The spheres on S5 at the S4 interface are the sites of mutations. Similar mutations on S4 involve deletions of the C-terminal helix, which is also at the S4-S5 interface. The other spheres on the S5 structure are the sites of resistance to spectinomycin.

10.1128/9781555818142/fig1-5_thmb.gif
10.1128/9781555818142/fig1-5.gif
Image of Figure 5
Figure 5

Importance of protein-protein interactions in the 30S subunit. The figure shows relative positions of proteins S4, S5, and S8 in the 30S structure. The spheres on S5 at the S4 interface are the sites of mutations. Similar mutations on S4 involve deletions of the C-terminal helix, which is also at the S4-S5 interface. The other spheres on the S5 structure are the sites of resistance to spectinomycin.

Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1
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Figure 6

Stereo view of our current interpretation of the 30S subunit at 5.5-Å resolution. The proteins of known structure are shown, along with S20, the central domain of 16S RNA, and H44. Regions that appear to be clearly double-stranded RNA in the map are shown in gray.

10.1128/9781555818142/fig1-6_thmb.gif
10.1128/9781555818142/fig1-6.gif
Image of Figure 6
Figure 6

Stereo view of our current interpretation of the 30S subunit at 5.5-Å resolution. The proteins of known structure are shown, along with S20, the central domain of 16S RNA, and H44. Regions that appear to be clearly double-stranded RNA in the map are shown in gray.

Citation: Ramakrishnan V, Capel M, Clemons, Jr. W, May J, Wimberly B. 2000. Progress toward the Crystal Structure of a Bacterial 30S Ribosomal Subunit, p 3-9. In Garett R, Douthwaite S, Liljas A, Matheson A, Moore P, Noller H (ed), The Ribosome. ASM Press, Washington, DC. doi: 10.1128/9781555818142.ch1
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