SEARCH · PubMed Health

Results for “rRNA structure”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 91 records · Page 5Linked to original sources

PubMed1973

Use of the 295- to 300-nanometer circular dichroism through of ribonucleic acid to study helix winding: effect of acridine orange.

Acridine orange decreases the amplitude of the 295-nm circular dichroism (CD) trough of ribosomal ribonucleic acid (rRNA) where the trough has been related to coil character. Since acridine orange is known from earlier work to intercalate between base pairs of nucleic acids, causing an unwinding of the coil, and our studies show a decrease in the 295-nm CD trough, it appears that CD measurements may be used to observe relative unwinding of rRNA. Under similar solution conditions, melting temperatures with acridine orange indicate no significant change in the stabilization of rRNA structure by acridine orange. Hypochromicity studies show no increase in the percent base pairing in rRNA when 0.1 M tris(hydroxymethyl)aminomethane (pH 7.6) with 1.35 M KCl is used. These results indicate that CD changes in the amplitude of the 295-nm trough of rRNA are related to helix winding in rRNA.

Acridines↗

PubMed2003

Conformational change in the 16S rRNA in the Escherichia coli 70S ribosome induced by P/P- and P/E-site tRNAPhe binding.

The effects of P/P- and P/E-site tRNA(Phe) binding on the 16S rRNA structure in the Escherichia coli 70S ribosome were investigated using UV cross-linking. The identity and frequency of 16S rRNA intramolecular cross-links were determined in the presence of deacyl-tRNA(Phe) or N-acetyl-Phe-tRNA(Phe) using poly(U) or an mRNA analogue containing a single Phe codon. For N-acetyl-Phe-tRNA(Phe) with either poly(U) or the mRNA analogue, the frequency of an intramolecular cross-link C967 x C1400 in the 16S rRNA was decreased in proportion to the binding stoichiometry of the tRNA. A proportional effect was true also for deacyl-tRNA(Phe) with poly(U), but the decrease in the C967 x C1400 frequency was less than the tRNA binding stoichiometry with the mRNA analogue. The inhibition of the C967 x C1400 cross-link was similar in buffers with, or without, polyamines. The exclusive participation of C967 with C1400 in the cross-link was confirmed by RNA sequencing. One intermolecular cross-link, 16S rRNA (C1400) to tRNA(Phe)(U33), was made with either poly(U) or the mRNA analogue. These results indicate a limited structural change in the small subunit around C967 and C1400 during tRNA P-site binding sensitive to the type of mRNA that is used. The absence of the C967 x C1400 cross-link in 70S ribosome complexes with tRNA is consistent with the 30S and 70S crystal structures, which contain tRNA or tRNA analogues; the occurrence of the cross-link indicates an alternative arrangement in this region in empty ribosomes.

Acetylation↗

PubMed1997

An unusual fibrillarin gene and protein: structure and functional implications.

The diploid germinal nucleus of the ciliated protozoan Tetrahymena thermophila is unusual among eukaryotes in that it encodes a single copy of the gene for rRNA allowing identification of cis-acting mutations in rDNA affecting rRNA structure, function, and processing. The generally conserved nucleolar protein fibrillarin has been characterized from a number of systems and is involved in pre-rRNA processing. We have demonstrated that Tetrahymena has fibrillarin and have analyzed the cDNA and the genomic DNA encoding this protein. The derived amino acid sequence of the N-terminal region of Tetrahymena fibrillarin shows little similarity with the generally highly conserved glycine/arginine-rich N-terminal domain of other eukaryotic fibrillarins. The remainder of the amino acid sequence of the molecule is more conserved. Polyclonal antibodies generated against the full-length Tetrahymena fibrillarin expressed in bacteria recognize a protein of M(r) approximately 32,000 in whole-cell or nucleolar preparations. Immunocytochemistry localizes fibrillarin to nucleoli in the somatic macronuclei of vegetative cells. Transformation experiments demonstrate that fibrillarin is an essential protein in Tetrahymena. The Tetrahymena fibrillarin is expressed but does not complement a NOP1 null mutation when transformed into the yeast Saccharomyces cerevisiae, indicating less functional conservation among fibrillarins than previously suggested.

Amino Acid Sequence↗

PubMed2000

Periodic conformational changes in rRNA: monitoring the dynamics of translating ribosomes.

In protein synthesis, a tRNA transits the ribosome via consecutive binding to the A (acceptor), P (peptidyl), and E (exit) site; these tRNA movements are catalyzed by elongation factor G (EF-G) and GTP. Site-specific Pb2+ cleavage was applied to trace tertiary alterations in tRNA and all rRNAs on pre- and posttranslocational ribosomes. The cleavage pattern of deacylated tRNA and AcPhe-tRNA changed individually upon binding to the ribosome; however, these different conformations were unaffected by translocation. On the other hand, translocation affects 23S rRNA structure. Significantly, the Pb2+ cleavage pattern near the peptidyl transferase center was different before and after translocation. This structural rearrangement emerged periodically during elongation, thus providing evidence for a dynamic and mobile role of 23S rRNA in translocation.

Base Sequence↗

PubMed1998

Assembly of proteins and 5 S rRNA to transcripts of the major structural domains of 23 S rRNA.

The six major structural domains of 23 S rRNA from Escherichia coli, and all combinations thereof, were synthesized as separate T7 transcripts and reconstituted with total 50 S subunit proteins. Analysis by one and two-dimensional gel electrophoresis demonstrated the presence of at least one primary binding protein associated with each RNA domain and additional proteins assembled to domains I, II, V and VI. For all the combinations of two to five domains, enhanced assembly yields and/or new proteins were observed primarily to those transcripts containing either domains I+II or domains V+VI. This indicates that there are two major protein assembly centres located at the ends of the 23 S rRNA, which is consistent with an earlier view that in vitro protein assembly nucleates around proteins L24 and L3. Although similar protein assembly patterns were observed over a range of temperature and magnesium concentrations, protein L2 assembled strongly with domains II and IV at 4-8 mM Mg2+ (the first step of the two-step reconstitution procedure) and with domain IV alone at higher Mg2+ concentrations (the second step). It is proposed that this change in protein-RNA binding provides a basis for the two-step reconstitution in vitro. A chemical footprinting approach was employed on the reconstituted protein-domain complexes to localize a putative L4 binding region within domain I to a region that is partially co-structural with the site on the L4-mRNA where L4 binds and inhibits its own translation. A similar approach was used to map the putative binding regions on domain V of protein L9 and the 5 S RNA-L5-L18 complex.

Base Sequence↗

PubMed1983

Consensus structure and evolution of 5S rRNA.

A consensus structure model of 5S rRNA presenting all conserved nucleotides in fixed positions has been deduced from the primary and secondary structure of 71 eubacterial, archaebacterial, eukaryotic cytosolic and organellar molecules. Phylogenetically related groups of molecules are characterized by nucleotide deletions in helices III, IV and V, and by potential base pair interactions in helix IV. The group-specific deletions are correlated with the early branching pattern of a dendrogram calculated from nucleotide substitution data: the first major division separates the group of eubacterial and organellar molecules from a second group containing the common ancestors of archaebacterial and eukaryotic/cytosolic molecules. The earliest diverging branch of the eubacterial/organellar group includes molecules from Thermus thermophilus, T. aquaticus, Rhodospirillum rubrum, Paracoccus denitrificans and wheat mitochondria.

Bacteria↗

PubMedDate not supplied

Structure of the rRNA genes in the hamster sperm nucleus.

We have examined the structure of the major ribosomal RNA (rRNA) genes in the hamster sperm nucleus, using fluorescent in situ hybridization (FISH). The rRNA genes are present as tandemly repeated clusters located at the telomeric ends of the short arms of five pairs of acrocentric chromosomes in the Syrian golden hamster (as they are in humans). In somatic cells, these five chromosome pairs come together to form the nucleolus, the site of rRNA synthesis. The nucleolus remains intact through S phase of the cell cycle, breaking apart only during late G2 and mitosis when the chromosomes condense. Mammalian sperm nuclei are the final products of meiotic division and morphological differentiation that includes a dramatic chromatin condensation. Consequently, it was not immediately obvious whether the rRNA genes would be condensed into a nucleolus-like structure in the mature spermatozoa, or separated, as they are in mitotic chromosomes. We found that of 117 sperm nuclei examined, 91.5% contained between two and five FISH signals for the rRNA gene clusters, and 64.0% contained four (29%) or five (35%) signals. In decondensed hamster sperm nuclei, the rRNA hybridized signals were separated into independent strands. These data collectively indicate that the chromosomes containing the rRNA genes are not bound together into a pre-nucleolar structure in fully condensed mammalian sperm nuclei.

Animals↗

PubMed2001

Structure of a eukaryotic decoding region A-site RNA.

The aminoglycoside antibiotics target a region of highly conserved nucleotides in the aminoacyl-tRNA site (A site) of 16 S RNA on the 30 S subunit. The structures of a prokaryotic decoding region A-site oligonucleotide free in solution and bound to the aminoglycosides paromomycin and gentamicin C1A have been determined. Here, the structure of a eukaryotic decoding region A-site oligonucleotide has been determined using homonuclear and heteronuclear NMR spectroscopy, and compared to the unbound prokaryotic rRNA structure. The two structures are similar, with a U1406-U1495 base-pair, a C1407-G1494 Watson-Crick base-pair, and a G1408-A1493 base-pair instead of the A1408-A1493 base-pair of the prokaryotic structure. The two structures differ in the orientation of the 1408 position with respect to A1493; G1408 is rotated toward the major groove, which is the binding pocket for aminoglycosides. The structures also differ in the stacking geometry of G1494 on A1493, which could have slight long-range conformational effects.

Base Pairing↗

PubMed1999

Distinct types of rRNA operons exist in the genome of the actinomycete Thermomonospora chromogena and evidence for horizontal transfer of an entire rRNA operon.

We describe here the presence of two distinct types of rRNA operons in the genome of a thermophilic actinomycete Thermomonospora chromogena. The genome of T. chromogena contains six rRNA operons (rrn), of which four complete and two incomplete ones were cloned and sequenced. Comparative analysis revealed that the operon rrnB exhibits high levels of sequence variations to the other five nearly identical ones throughout the entire length of the operon. The coding sequences for the 16S and 23S rRNA genes differ by approximately 6 and 10%, respectively, between the two types of operons. Normal functionality of rrnB is concluded on the basis of the nonrandom distribution of nucleotide substitutions, the presence of compensating nucleotide covariations, the preservation of secondary and tertiary rRNA structures, and the detection of correctly processed rRNAs in the cell. Comparative sequence analysis also revealed a close evolutionary relationship between rrnB operon of T. chromogena and rrnA operon of another thermophilic actinomycete Thermobispora bispora. We propose that T. chromogena acquired rrnB operon from T. bispora or a related organism via horizontal gene transfer.

Actinomycetales↗

PubMed1987

A 5 S rRNA-like secondary structure in the 7 SL RNA may define a ribosomal binding site of the signal recognition particle.

A new secondary structure model for parts of the 7 SL RNA is proposed which indicates for a stretch of at least 40 bases a strong structural homology to the ribosomal protein L5 binding site of eukaryotic 5 S rRNA. It is suggested that the 5 S rRNA-like structural part of 7 SL RNA mediates binding of the signal recognition particle near to the peptidyl transferase center of the ribosome.

Animals↗

PubMed1995

Comparison of Mycobacterium 23S rRNA sequences by high-temperature reverse transcription and PCR.

We describe a modified rRNA sequence analysis method which we used to determine the phylogenetic relationships among 58 species belonging to the genus Mycobacterium. We combined the sensitivity of the reverse transcriptase PCR for amplifying nanogram amounts of template rRNA material with the elevated extension temperatures used for the thermostable DNA polymerase from Thermus thermophilus. A 70 degrees C reverse transcription extension step permitted improved read-through of highly structured rRNA templates from members of the genus Mycobacterium, which have G+C contents of 66 to 71 mol%. The nucleic acid sequences of the amplified material were then determined by performing thermal cycle sequencing with alpha-33P-labeled primers, again with extension at 70 degrees C. Nonspecifically terminated bands were chased by using terminal deoxynucleotidyl transferase. Our method had a template requirement of nanogram amounts or less of purified RNA or 2,000 CFU of intact cells and had sufficient sensitivity so that lyophils obtained from the American Type Culture Collection could be used as source material. Sequences from a 250-nucleotide stretch of the 23S rRNA were aligned, and phylogenetic trees were evaluated by using the De Soete distance treeing algorithm and Rhodococcus bronchialis as the outgroup. Our 23S rRNA trees were compared with previously published 16S rRNA trees, including the comprehensive trees developed by the University of Illinois Ribosomal Database Project, and included 15 species not evaluated previously. Most of the groups were in general agreement and were consistent with relationships determined on the basis of biochemical characteristics, but some new relationships were also observed.

Base Sequence↗

PubMed1982

Allele-specific, selective amplification of a ribosomal RNA gene in Tetrahymena thermophila.

The amplification of ribosomal DNA during development of the somatic macronucleus in Tetrahymena thermophila was analyzed by genetic and molecular biological techniques. We have identified an alternate form of the rDNA, structurally distinguishable from the wild-type by an extra cutting site for Bam HI in its nontranscribed spacer. The altered rDNA was inherited in crosses in a simple Mendelian fashion, consistent with the presence of only one rRNA gene copy per haploid genome in the micronucleus. We therefore define a locus for the rRNA structural gene, the rdnA locus, with the allele determining the alternate form designated rdnA1. In over 95% of T. thermophila clones heterozygous for the rdnA locus in the micronucleus (rdnA1/rdn+), the macronucleus, which develops from a division product of this micronucleus, contained almost exclusively rdnA1-type amplified palindromic rDNA molecules. The rdnA1 allele is thus almost always dominant over the rdn+ allele with respect to amplification. This genetic variant of the rdnA locus was used to show that the single, free, nonpalindromic rRNA genes, which are synthesized during rDNA amplification, are derived from micronuclear gene copies from both chromosomal homologs. We therefore conclude that in these heterozygotes, selective amplification of the rdnA1 allele is not caused by the production of only one type of free, single rRNA gene during amplification.

Alleles↗

PubMed1993

Physiological and genetic regulation of rRNA synthesis in Lactococcus.

The macromolecular composition of Lactococcus was regulated by growth rate in the same general way as that of less fastidious bacteria such as Escherichia coli and Salmonella typhimurium. The ratios of RNA:DNA and RNA:protein increased approximately threefold over a 13.5-fold increase in growth rate, whereas the ratio of DNA:protein remained approximately constant. Using reporter genes fused to a DNA fragment of a cloned lactococcal rRNA operon, promoter activity was located upstream of the 16S rRNA structural gene. This DNA fragment had some characteristics typical of a rrn promoter in E. coli. Two consensus promoter sequences P1 and P2 were located 296 and 157 bp, respectively, upstream of the start of the 16S rRNA gene. Between P2 and the start of the 16S rRNA gene, sequences were identified with typical anti-termination motifs characteristic of E. coli rrn promoter regions. A putative transcription terminator sequence was identified downstream of the 5S rRNA gene and putative primary RNA transcript processing sites at both ends of the lactococcal rRNA operon were also noted.

Amino Acid Sequence↗

PubMed1996

Conserved sequence motifs, alignment, and secondary structure for the third domain of animal 12S rRNA.

Secondary structure models are an important step for aligning sequences, understanding probabilities of nucleotide substitutions, and evaluating the reliability of phylogenetic reconstructions. A set of conserved sequence motifs is derived from comparative sequence analysis of 184 invertebrate and vertebrate taxa (including many taxa from the same genera, families, and orders) with reference to a secondary structure model for domain III of animal mitochondrial small subunit (12S) ribosomal RNA. A template is presented to assist with secondary structure drawing. Our model is similar to previous models but is more specific to mitochondrial DNA, fitting both invertebrate and vertebrate groups, including taxa with markedly different nucleotide compositions. The second half of the domain III sequence can be difficult to align precisely, even when secondary structure information is considered. This is especially true for comparisons of anciently diverged taxa, but well-conserved motifs assist in determining biologically meaningful alignments. Patterns of conservation and variability in both paired and unpaired regions make differential phylogenetic weighting in terms of "stems" and "loops" unsatisfactory. We emphasize looking carefully at the sequence data before and during analyses, and advocate the use of conserved motifs and other secondary structure information for assessing sequencing fidelity.

Animals↗

PubMed2005

High-level chromosomally mediated tetracycline resistance in Neisseria gonorrhoeae results from a point mutation in the rpsJ gene encoding ribosomal protein S10 in combination with the mtrR and penB resistance determinants.

Neisseria gonorrhoeae becomes resistant to tetracycline by two major mechanisms: expression of a plasmid-encoded TetM protein and mutations in endogenous genes (chromosomally mediated resistance). Early studies by Sparling and colleagues (P. F. Sparling F. A. J. Sarubbi, and E. Blackman, J. Bacteriol. 124:740-749, 1975) demonstrated that three genes were involved in high-level chromosomally mediated tetracycline resistance (MIC of tetracycline > or = 2 microg/ml): ery-2 (now referred to as mtrR), penB, and tet-2. While the identities of the first two genes are known, the tet-2 gene has not been identified. We cloned the tet-2 gene, which confers tetracycline resistance, from tetracycline-resistant clinical isolate N. gonorrhoeae FA6140 and show that resistance is due to a single point mutation (Val-57 to Met) in the rpsJ gene (rpsJ1) encoding ribosomal protein S10. Moreover, the identical mutation was found in six distinct tetracycline-resistant clinical isolates in which the MIC of tetracycline was > or =2 microg/ml. Site-saturation mutagenesis of the codon for Val-57 identified two other amino acids (Leu and Gln) that conferred identical levels of resistance as the Met-57 mutation. The mutation maps to the vertex of a loop in S10 that is near the aminoacyl-tRNA site in the structure of the 30S ribosomal subunit from Thermus thermophilus, and the residue equivalent to Val-57 in T. thermophilus S10, Lys-55, is within 8 to 9 A of bound tetracycline. These data suggest that large noncharged amino acids alter the rRNA structure near the tetracycline-binding site, leading to a lower affinity of the antibiotic.

Alleles↗

PubMed2005

Graphical representation of ribosomal RNA probe accessibility data using ARB software package.

BACKGROUND: Taxon specific hybridization probes in combination with a variety of commonly used hybridization formats nowadays are standard tools in microbial identification. A frequently applied technology, fluorescence in situ hybridization (FISH), besides single cell identification, allows the localization and functional studies of the microbial community composition. Careful in silico design and evaluation of potential oligonucleotide probe targets is therefore crucial for performing successful hybridization experiments. RESULTS: The PROBE Design tools of the ARB software package take into consideration several criteria such as number, position and quality of diagnostic sequence differences while designing oligonucleotide probes. Additionally, new visualization tools were developed to enable the user to easily examine further sequence associated criteria such as higher order structure, conservation, G+C content, transition-transversion profiles and in situ target accessibility patterns. The different types of sequence associated information (SAI) can be visualized by user defined background colors within the ARB primary and secondary structure editors as well as in the PROBE Match tool. CONCLUSION: Using this tool, in silico probe design and evaluation can be performed with respect to in situ probe accessibility data. The evaluation of proposed probe targets with respect to higher-order rRNA structure is of importance for successful design and performance of in situ hybridization experiments. The entire ARB software package along with the probe accessibility data is available from the ARB home page http://www.arb-home.de

Algorithms↗

PubMed1983

Complementary DNA-25S ribosomal RNA hybridization: an improved method for phylogenetic studies.

In a new combination of techniques for ribosomal RNA hybridization, complementary DNA is synthesized on 25S ribosomal RNA fragments generated by mild alkali treatment, by the enzymatic addition of polyadenylic acid tails, hybridization of these tails with oligo deoxyribosylthymine, and reverse transcription in the presence of tritiated TTP. Hybridization reactions are performed in solution. Heteroduplexes are collected on diethylaminoethylcellulose filter discs after treatment with S1 nuclease. The problems presented by secondary rRNA structure are avoided by denaturation before reverse transcription and before hybridization. The high percentage of duplex formation (78-87%), the low standard deviation of relative binding (averaging +/- 1.30% relative binding), and small differences in reciprocal hybridizations (1.71-5.18% relative binding), as well as the elimination of complications resulting from differences in the number of rRNA cistrons in nuclear DNA, make this method preferable to the membrane-filter technique commonly used in phylogenetic classifications based on the homology of large rRNAs.

DNA↗

PubMed1993

The lepidopteran mitochondrial control region: structure and evolution.

For several species of lepidoptera, most of the approximately 350-bp mitochondrial control-region sequences were determined. Six of these species are in one genus, Jalmenus; are closely related; and are believed to have undergone recent rapid speciation. Recent speciation was supported by the observation of low interspecific sequence divergence. Thus, no useful phylogeny could be constructed for the genus. Despite a surprising conservation of control-region length, there was little conservation of primary sequences either among the three lepidopteran genera or between lepidoptera and Drosophila. Analysis of secondary structure indicated only one possible feature in common--inferred stem loops with higher-than-random folding energies--although the positions of the structures in different species were unrelated to regions of primary sequence similarity. We suggest that the conserved, short length of control regions is related to the observed lack of heteroplasmy in lepidopteran mitochondrial genomes. In addition, determination of flanking sequences for one Jalmenus species indicated (i) only weak support for the available model of insect 12S rRNA structure and (ii) that tRNA translocation is a frequent event in the evolution of insect mitochondrial genomes.

Animals↗