Activities of <i>B. stearothermophilus</i> 50 S ribosomes reconstituted with prokaryotic and eukaryotic 5 S RNA

Wrede, Paul; Erdmann, Volker A.

doi:10.1016/0014-5793(73)80219-4

Cited by 68 publications

(28 citation statements)

References 24 publications

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“…This extreme ribosomal complexity is believed to be the basis for the species specificity in 16S rRNA, although solid experimental evidence is not available in this regard (4,6). It should be noted that 5S rRNA genes, which have less complex structures, are known to be freely exchangeable in bacteria, at least in vitro (25). Using E. coli as a host, Fox's group has also shown that 5S rRNA from any Vibrio species, except for Vibrio gazogenes, could be horizontally transferred (26).…”

Section: Resultsmentioning

confidence: 99%

Mutational robustness of 16S ribosomal RNA, shown by experimental horizontal gene transfer in Escherichia coli

Kitahara

Yasutake

Miyazaki

2012

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

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The bacterial ribosome consists of three rRNA molecules and 57 proteins and plays a crucial role in translating mRNA-encoded information into proteins. Because of the ribosome's structural and mechanistic complexity, it is believed that each ribosomal component coevolves to maintain its function. Unlike 5S rRNA, 16S and 23S rRNAs appear to lack mutational robustness, because they form the structural core of the ribosome. However, using Escherichia coli Δ7 (null mutant of operons) as a host, we have recently shown that an active hybrid ribosome whose 16S rRNA has been specifically substituted with that from non-E. coli bacteria can be reconstituted in vivo. To investigate the mutational robustness of 16S rRNA and the structural basis for its functionality, we used a metagenomic approach to screen for 16S rRNA genes that complement the growth of E. coli Δ7. Various functional genes were obtained from the Gammaproteobacteria and Betaproteobacteria lineages. Despite the large sequence diversity (80.9-99.0% identity with E. coli 16S rRNA) of the functional 16S rRNA molecules, the doubling times (DTs) of each mutant increased only modestly with decreasing sequence identity (average increase in DT, 4.6 s per mutation). The three-dimensional structure of the 30S ribosome showed that at least 40.7% (628/1,542) of the nucleotides were variable, even at ribosomal protein-binding sites, provided that the secondary structures were properly conserved. Our results clearly demonstrate that 16S rRNA functionality largely depends on the secondary structure but not on the sequence itself.functional plasticity T he ribosome plays a crucial role in translating mRNA-encoded information into proteins, which consists of 2 (large and small) subunits. In prokaryotes, the small 30S subunit consists of 16S rRNA and 21 proteins, whereas the large 50S subunit consists of 23S rRNA, 5S rRNA, and 36 proteins. Both in bacteria and archaea, 16S or 23S rRNA shapes the structural core of the subunit particle, whereas many ribosomal proteins are often found on the surface of the subunit, with extensions that protrude into the RNA core (1-3). In contrast, 5S rRNA (small RNA with 120 bases) simply attaches to the 50S subunit and constitutes a structure called a central protuberance

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Section: Resultsmentioning

confidence: 99%

Mutational robustness of 16S ribosomal RNA, shown by experimental horizontal gene transfer in Escherichia coli

Kitahara

Yasutake

Miyazaki

2012

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

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“…The 5-S RNA from H. cutiruhrum will bind BL5 and BL22. the equivalent proteins in B. stearotliermophilus and the resulting complex can be incorporated into B. stearotlzernzophilus 50-S subunits to yield active ribosomes [28] and (Wrede, Matheson and Erdmann, unpublished results). Work is now in progress on the sequence of HL13, HL19 and 5-S RNA from the extreme halophile.…”

Section: Discussionmentioning

confidence: 99%

The 5‐S RNA · Protein Complex from an Extreme Halophile, Halobacterium cutirubrum

Smith

Yaguchi

Willick

et al. 1978

European Journal of Biochemistry

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A 5-S RNA . protein complex has been isolated from the 50-S ribosomal subunit of an extreme halophile, Hulobacterium cutirubrum. The 50-S ribosomal subunit from the extreme halophile requires 3.4 M K' and 100 mM Mg" for stability. However, if the high K' concentration is maintained but the Mg2' concentration lowered to 0.3 mM, the 5-S RNA . protein complex is selectively extracted from the subunit. After being purified on an Agarose 0.5-m column the complex had a molecular weight of about 80000 and contained 5-S RNA and two proteins, HL13 and HL19, with molecular weights (by sedimentation equilibrium) of 18 700 and 18000, respectively. No ATPase or GTPase activity could be detected in the 5-S RNA . protein complex. The amino acid composition and electrophoretic mobility on polyacrylamidc gels indicated both proteins were much more acidic than the equivalent from Escherichia coli or Bacillus steurotliermoI)hilus. Partial amino acid sequence data suggest HL13 is homologous to EL18 and HL19 to EL5.The 5-S RNA, a component of the 50-S ribosomal subunit of all prokaryotes, is believed to play a key role in the structure and function of the ribosome. Studies by Erdmann and his co-workers (see [1] for review) have indicated that the 5-S RNA forms part of the aminoacyl-tRNA binding site on the ribosome specifically interacting with the T-Y-C-G sequence in the tRNA through a G-A-A-C nucleotide sequence. The 5-S RNA is also thought to be involved in the translocation step in protein synthesis [2].In Eschericliiu coli, two proteins, ELI8 and EL25, specifically bind to the 5-S RNA [3] while in the therniophile Bacillus stcarotlicrrnc~~~hilus BL5 and BL22 have a similar role [3]. There is also evidence in E. coli that a third protein EL5 is also part of the 5-S RNA . protein complex [4]. The sequence of the 5-S RNA from a large number of prokaryotes has been determined [5] and the complete amino acid sequences of the ribosomal proteins EL5 [ 6 ] , EL18 [7] and EL25 [8,9] are now known.

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“…It was generally believed that the chances of an organism being able to use rRNA from any other organism are very low but there are evidences that functional ribosomes can be reconstituted from different organisms [57][58][59][60] . Each ribosomal operon of an organism can be replaced with those of another species 61,62 and divergent rRNA operons can coexist in the same genome 63,64 .…”

Section: Pitfalls Of Polyphasic Approachmentioning

confidence: 99%

Polyphasic approach of bacterial classification — An overview of recent advances

et al. 2007

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Classification of microorganisms on the basis of fi traditional microbiological methods (morphological, physiological and biochemical) creates a blurred image about their taxonomic status and thus needs further clarification. It fi should be based on a more pragmatic approach of deploying a number of methods for the complete characterization of microbes. Hence, the methods now employed for bacterial systematics include, the complete 16S rRNA gene sequencing and its comparative analysis by phylogenetic trees, DNA-DNA hybridization studies with related organisms, analyses of molecular markers and signature pattern(s), biochemical assays, physiological and morphological tests. Collectively these genotypic, chemotaxonomic and phenotypic methods for determining taxonomic position of microbes constitute what is known as the 'polyphasic approach' for bacterial systematics. This approach is currently the most popular choice for classifying bacteria and several microbes, which were previously placed under invalid taxa have now been resolved into new genera and species. This has been possible owing to rapid development in molecular biological techniques, automation of DNA sequencing coupled with advances in bioinformatic tools and access to sequence databases. Several DNA-based typing methods are known; these provide information for delineating bacteria into different genera and species and have the potential to resolve differences among the strains of a species. Therefore, newly isolated strains must be classifi ed fi on the basis of the polyphasic approach. Also previously classifi ed organisms, as and when required, can be reclasfi sified on this ground in order to obtain information about fi their accurate position in the microbial world. Thus, current techniques enable microbiologists to decipher the natural phylogenetic relationships between microbes.

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Activities of B. stearothermophilus 50 S ribosomes reconstituted with prokaryotic and eukaryotic 5 S RNA

Cited by 68 publications

References 24 publications

Mutational robustness of 16S ribosomal RNA, shown by experimental horizontal gene transfer in Escherichia coli

Mutational robustness of 16S ribosomal RNA, shown by experimental horizontal gene transfer in Escherichia coli

The 5‐S RNA · Protein Complex from an Extreme Halophile, Halobacterium cutirubrum

Polyphasic approach of bacterial classification — An overview of recent advances

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