Structure and Interactions of Ribosomal Components and Ligands

Ebel, Jean‐Pierre; Ehresmann, Bernard; Ehresmann, Chantal; Mougel, Marylène; Giegé, R.; Moras, Dino

doi:10.1007/978-1-4612-4884-2_16

Cited by 3 publications

(2 citation statements)

References 44 publications

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“…However, it is possible that S6, like S16 or S18, plays an important role in the assembly process, which may be masked because the presence of all of the other proteins results in cooperative interactions leading to the formation of particles apparently similar to the native subunits (32). Secondary structure models have delineated the 16S rRNA into three major domains (5', central, and 3') held together by a pseudoknot structure, which forms a "topographical center" (24,(33)(34)(35). The mass centers observed at this stage of the assembly process are analogous to the three major domains ofthe 16S rRNA, which are held together by the topographical center.…”

Section: Resultsmentioning

confidence: 99%

Assembly of the Escherichia coli 30S ribosomal subunit reveals protein-dependent folding of the 16S rRNA domains.

Mandiyan

Tumminia

Wall

et al. 1991

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

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Protein-nucleic acid interactions involved in the assembly process of the Escherichia coli 30S ribosomal subunit were quantitatively analyzed by high-resolution scanning transmission electron microscopy. The in vitro reconstituted ribonucleoprotein (core) particles were characterized by their morphology, mass, and radii of gyration. During the assembly of the 30S subunit, the 16S rRNA underwent significant conformational changes that were governed by the cooperative interactions of the ribosomal proteins. The sequential association of the first 12 proteins with the 16S rRNA resulted in the formation of core particles containing up to three mass centers at distinct stages of the assembly process. These globular mass centers may correspond to the three major domains (5', central, and 3') of the 16S rRNA. Through the subsequent interactions of the late assembly proteins with the 16S rRNA, two of the three domains merge, yielding the basic structural traits ofthe native 30S subunit. The fine morphological features of the native 30S subunit became distinctly resolved only after the addition of the full complement of proteins. The fully reconstituted 30S subunits are active in polyphenylalanine synthesis assays. Visualiztion of the assembly mechanism of the E. coli 30S ribosomal subunit revealed domain-specific folding of the 16S rRNA through the formation bf distinct intermediate core particles hitherto not observed.Ribosomes are ubiquitous macromolecular assemblies of proteins and ribonucleic acids involved in protein biosynthesis. This bipartite cellular organelle is composed of a large and a small subunit. The structure-function relationship of the small (30S) ribosomal subunit ofEscherichia coli has been extensively studied. This subunit is actively involved in the initiation of protein synthesis by its interactions with initiation factors, messenger RNAs, transfer RNAs, and the large (50S) ribosomal subunit (1-4). The 30S subunit possesses characteristic structural features known as the head, body, cleft, and platform (5, 6) and is composed of a single 16S rRNA molecule and 21 different ribosomal proteins (S1-S21). Several ribosomal proteins interact with the 16S rRNA through a sequential and cooperative process as revealed from the in vitro assembly of the 30S subunit (7-9). Results obtained from these studies led to the classification of the proteins as primary binding proteins, which bind directly to the 16S rRNA, and secondary binding proteins, which require the previous association of the primary binding proteins. Apart from being the structural base for protein interactions, the 16S rRNA has an established role in protein biosynthesis (10-14). However, the role of the ribosomal proteins in ribosome structure and function remains unclear.Scanning transmission electron microscopy (STEM) was used to quantitatively evaluate the conformational changes induced in the 16S rRNA molecule by its interactions with the ribosomal proteins under nondenaturing conditions (15). Unlike other physical techniques tha...

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Section: Resultsmentioning

confidence: 99%

Assembly of the Escherichia coli 30S ribosomal subunit reveals protein-dependent folding of the 16S rRNA domains.

Mandiyan

Tumminia

Wall

et al. 1991

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

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“…The information used to compose the complex secondary structure models for the 16S rRNA molecule has mostly been derived from chemical, enzymatic and phylogenetic data enIRL Press Limited, Oxford, England (reviewed in Woese et al, 1983;Brimacombe, 1985;Ebel et al, 1985;Lake, 1985). Mutational analyses have so far not contributed to the assembly and verification of this model because mutations in the 16S rRNA structural gene are relatively rare and limited to mutations conferring antibiotic resistance (Sigmund et al, 1984;Noller, 1986, 1987;and reviewed in Noller, 1984).…”

Section: Introductionmentioning

confidence: 99%

Mutagenesis at the mRNA decoding site in the 16S ribosomal RNA using the specialized ribosome system in Escherichia coli.

Hui¹,

Eaton²,

Boer³

1988

The EMBO Journal

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In the specialized ribosome system, a distinct pool of mutated ribosomes is dedicated to the translation of one particular mRNA species. This was accomplished by altering the Shine-LDalgarno sequence on the mRNA and its complementary anti-Shine -Dalgarno sequence on the plasmid-borne 16S rRNA gene. Here, using the specialized ribosome system, we were able to introduce mutations in key regions of the 16S rRNA and could study their effect on translation in vivo. The C 1400 region has been implicated to play a role in the actual mRNA decoding process. Several ribosomal mutations were introduced in this region. We showed that substitution of the evolutionary highly conserved C1400 residue by a G-or an A-residue inhibits ribosomal activity by 80% and 50% respectively, whereas, a C to a U change at this conserved position does not affect overall ribosomal activity. The adjacent stem structure (1410-1490) was also examined. Disruption of the stem by replacing either one of the arms of this stem, with a different sequence, inhibits ribosomal activity by -80%. A small but significant restoration of translation could be achieved by recreating a complementary stem with a different sequence. We found that full reversion of activity could be obtained when such mutated ribosomes were made spectinomycin resistant by introducing a C to A substitution at position 1192 which is located far away in the secondary structure map of the 16S rRNA molecule. Based on these results we conclude that some, but not all, of the nucleotides in the conserved C1400 region play a key role in translation. Most importantly, we show that the specialized ribosome system provides a very powerful tool for structure and function studies of any region in the complex ribosomal RNA molecule.

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A Genetic Approach to Molecular Interactions in the Yeast Mitochondrial Ribosome

Daignan‐Fornier¹,

Su²,

Contamine³

et al. 1988

Genetics of Translation

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Structure and Interactions of Ribosomal Components and Ligands

Cited by 3 publications

References 44 publications

Assembly of the Escherichia coli 30S ribosomal subunit reveals protein-dependent folding of the 16S rRNA domains.

Assembly of the Escherichia coli 30S ribosomal subunit reveals protein-dependent folding of the 16S rRNA domains.

Mutagenesis at the mRNA decoding site in the 16S ribosomal RNA using the specialized ribosome system in Escherichia coli.

A Genetic Approach to Molecular Interactions in the Yeast Mitochondrial Ribosome

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