An oligonucleotide analog approach to the decoding region of 16S rRNA

1995 ◽  
Vol 73 (11-12) ◽  
pp. 899-905 ◽  
Author(s):  
Seth Stern ◽  
Prakash Purohit
Keyword(s):  
16S Rrna ◽  
Ribosomal Rna ◽  
Molecular Basis ◽  
Ribosomal Subunit ◽  
Small Subunit ◽  
16S Ribosomal Rna ◽  
New Approach ◽  
16S Rna ◽  
Analog Approach ◽  
Decoding Region

Despite the passage of about 30 years since the discovery of the translational activities of ribosomes and the outlining of the roles of the large and small subunits, the actual molecular basis for the mRNA decoding activities of the small subunit has remained essentially obscure. In this paper, we describe a new approach using oligonucleotide analogs of 16S ribosomal RNA, in which the small ribosomal subunit is effectively deconstructed into a smaller more experimentally tractable form. Specifically, we review the results of experiments using an oligonucleotide analog of the decoding region of 16S ribosomal RNA, suggesting that the decoding region is the functional core of the small subunit, that it contacts both mRNA codons and tRNA anticodons, and that it mediates and probably enhances codon–anticodon base pairing, that is, decoding.Key words: translation, ribosome, 30S, 16S, RNA, decoding, antibiotic.

Protein-induced conformational changes in 16S rRNA during assembly of the Escherichia Coli 30S ribosomal subunit: A STEM study

1987 ◽  
Vol 45 ◽  
pp. 910-911
Author(s):  
M. Boublik ◽  
V. Mandiyan ◽  
J.F. Hainfeld ◽  
J.S. Wall
Keyword(s):  
16S Rrna ◽  
Conformational Changes ◽  
Ribosomal Proteins ◽  
Structural Changes ◽  
Ribosomal Subunit ◽  
Compact Structure ◽  
E Coli ◽  
Freeze Dried ◽  
16S Rna ◽  
30S Subunit

The aim of this study is to understand the mechanism of 16S rRNA folding into the compact structure of the small 30S subunit of E. coli ribosome. The assembly of the 30S E. coli ribosomal subunit is a sequence of specific interactions of 16S rRNA with 21 ribosomal proteins (S1-S21). Using dedicated high resolution STEM we have monitored structural changes induced in 16S rRNA by the proteins S4, S8, S15 and S20 which are involved in the initial steps of 30S subunit assembly. S4 is the first protein to bind directly and stoichiometrically to 16S rRNA. Direct binding also occurs individually between 16S RNA and S8 and S15. However, binding of S20 requires the presence of S4 and S8. The RNA-protein complexes are prepared by the standard reconstitution procedure, dialyzed against 60 mM KCl, 2 mM Mg(OAc)2, 10 mM-Hepes-KOH pH 7.5 (Buffer A), freeze-dried and observed unstained in dark field at -160°.

scholarly journals Evaluation of 16S, map1 and pCS20 probes for detection of Cowdria and Ehrlichia species

1999 ◽  
Vol 122 (2) ◽  
pp. 323-328 ◽  
Author(s):  
M. T. E. P. ALLSOPP ◽  
C. M. HATTINGH ◽  
S. W. VOGEL ◽  
B. A. ALLSOPP
Keyword(s):  
16S Rrna ◽  
Ribosomal Rna ◽  
Pcr Amplification ◽  
16S Ribosomal Rna ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Amblyomma Hebraeum ◽  
Ribosomal Rna Gene ◽  
Group Iii ◽  
16S Ribosomal Rna Gene

A panel of 16S ribosomal RNA gene probes has been developed for the study of the epidemiology of heartwater; five of these detect different cowdria genotypes, one detects five distinct genotypes; one detects any Group III Ehrlichia species other than Cowdria and one detects any Group II Ehrlichia species. These probes have been used on PCR-amplified rickettsial 16S rRNA genes from over 200 Amblyomma hebraeum ticks. Control ticks were laboratory-reared and either uninfected or fed on sheep experimentally infected with different cowdria isolates, field ticks were collected from animals in heartwater-endemic areas. All tick-derived DNA samples were also examined by PCR amplification and probing for two other cowdria genes (map1 and pCS20) which have previously been used for heartwater epidemiology. This paper describes the first direct comparison of all currently available DNA probes for heartwater-associated organisms.

scholarly journals Detection of Nocardia by 16S Ribosomal RNA Gene PCR and Metagenomic Next-Generation Sequencing (mNGS)

2022 ◽  
Vol 11 ◽  
Author(s):  
Juanjuan Ding ◽  
Bing Ma ◽  
Xupeng Wei ◽  
Ying Li
Keyword(s):  
16S Rrna ◽  
Ribosomal Rna ◽  
Human Herpesvirus ◽  
Antibiotic Sensitivity ◽  
Common Species ◽  
16S Ribosomal Rna ◽  
Clinical Samples ◽  
Klebsiella Pneumonia ◽  
Antimicrobial Drugs ◽  
Detection And Identification

In this study, the aim was to investigate the discriminatory power of molecular diagnostics based on mNGS and traditional 16S ribosomal RNA PCR among Nocardia species. A total of fourteen clinical isolates from patients with positive Nocardia cultures and clinical evidence were included between January 2017 and June 2020 in HeNan Provincial People’s Hospital. DNA extraction and 16S rRNA PCR were performed on positive cultures, and pathogens were detected by mNGS in these same samples directly. Among the 14 Nocardia isolates, four species were identified, and N. cyriacigeorgica (8 cases) is the most common species. Twelve of the 14 Nocardia spp. isolates were identified by the two methods, while two strains of N. cyriacigeorgica were not identified by mNGS. All tested isolates showed susceptibility to trimethoprim-sulfamethoxazole (SXT), amikacin and linezolid. Apart from Nocardia species, other pathogens such as Acinetobacter baumannii, Klebsiella pneumonia, Aspergillus, Enterococcus faecalis, Human herpesvirus, etc., were detected from the same clinical samples by mNGS. However, these different pathogens were considered as colonization or contamination. We found that it is essential to accurately identify species for determining antibiotic sensitivity and, consequently, choosing antibiotic treatment. 16S rRNA PCR was useful for identification of nocardial infection among species, while this technique needs the clinicians to make the pre-considerations of nocardiosis. However, mNGS may be a putative tool for rapid and accurate detection and identification of Nocardia, beneficial for applications of antimicrobial drugs and timely adjustments of medication.

Heterogeneity between 16S ribosomal RNA gene copies borne by one Desulfitobacterium strain is caused by different 100-200 bp insertions in the 5´ region

2007 ◽  
Vol 53 (1) ◽  
pp. 116-128 ◽  
Author(s):  
Richard Villemur ◽  
Philippe Constant ◽  
Annie Gauthier ◽  
Martine Shareck ◽  
Réjean Beaudet
Keyword(s):  
In Situ Hybridization ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Ribosomal Rna ◽  
16S Ribosomal Rna ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Gene Copies

Strains of Desulfitobacterium hafniense, such as strains PCP-1, DP7, TCE1, and TCP-A, have unusual long 16S ribosomal RNA (rRNA) genes due to an insertion of approximately 100 bp in the 5' region. In this report, we analyzed the 16S rRNA genes of different Desulfitobacterium strains to determine if such an insertion is a common feature of desulfitobacteria. We amplified this region by polymerase chain reaction (PCR) from eight Desulfitobacterium strains (D. hafniense strains PCP-1, DP7, TCP-A, TCE1, and DCB-2; D. dehalogenans; D. chlororespirans; and Desulfitobacterium sp. PCE1) and resolved each PCR product by denaturing gradient gel electrophoresis (DGGE). All strains had from two to seven DGGE- migrating bands, suggesting heterogeneity in their 16S rRNA gene copies. For each strain, the 5' region of the 16S rRNA genes was amplified and a clone library was derived. Clones corresponding to most PCR–DGGE migration bands were isolated. Sequencing of representative clones revealed that the heterogeneity was generated by insertions of 100–200 bp. An insertion was found in at least one copy of the 16S rRNA gene in all examined strains. In total, we found eight different types of insertions (INS1–INS8) that varied from 123 to 193 nt in length. Two-dimensional structural analyses of transcribed sequences predicted that all insertions would form an energetically stable loop. Reverse transcriptase – PCR experiments revealed that most of the observed insertions in the Desulfitobacterium strains were excised from the mature 16S rRNA transcripts. Insertions were not commonly found in bacterial 16S rRNA genes, and having a different insertion in several 16S rRNA gene copies borne by a single bacterial species was rarely observed. The function of these insertions is not known, but their occurrence can have an important impact in deriving 16S rRNA oligonucleotidic fluorescence in situ hybridization probes, as these insertions can be excised from 16S rRNA transcripts.Key words: Desulfitobacterium, 16S ribosomal RNA genes, heterogeneity, gene insertions, fluorescence in situ hybridization.

Conformational changes in 16S ribosomal RNA induced by 30S ribosomal subunit proteins from Escherichia coli

Science ◽  
1978 ◽  
Vol 202 (4371) ◽  
pp. 999-1001 ◽  
Author(s):  
A. Bogdanov ◽  
R. Zimmermann ◽  
C. Wang ◽  
N. Ford
Keyword(s):  
Escherichia Coli ◽  
Ribosomal Rna ◽  
Conformational Changes ◽  
Ribosomal Subunit ◽  
16S Ribosomal Rna

scholarly journals Human ERAL1 is a mitochondrial RNA chaperone involved in the assembly of the 28S small mitochondrial ribosomal subunit

2010 ◽  
Vol 430 (3) ◽  
pp. 551-558 ◽  
Author(s):  
Sven Dennerlein ◽  
Agata Rozanska ◽  
Mateusz Wydro ◽  
Zofia M. A. Chrzanowska-Lightowlers ◽  
Robert N. Lightowlers
Keyword(s):  
16S Rrna ◽  
Cellular Localization ◽  
Mitochondrial Protein ◽  
Ribosomal Subunit ◽  
Small Subunit ◽  
Mitochondrial Rna ◽  
Rna Chaperone ◽  
Stem Loop ◽  
Loop Region

The bacterial Ras-like protein Era has been reported previously to bind 16S rRNA within the 30S ribosomal subunit and to play a crucial role in ribosome assembly. An orthologue of this essential GTPase ERAL1 (Era G-protein-like 1) exists in higher eukaryotes and although its exact molecular function and cellular localization is unknown, its absence has been linked to apoptosis. In the present study we show that human ERAL1 is a mitochondrial protein important for the formation of the 28S small mitoribosomal subunit. We also show that ERAL1 binds in vivo to the rRNA component of the small subunit [12S mt (mitochondrial)-rRNA]. Bacterial Era associates with a 3′ unstructured nonanucleotide immediately downstream of the terminal stem–loop (helix 45) of 16S rRNA. This site contains an AUCA sequence highly conserved across all domains of life, immediately upstream of the anti-Shine–Dalgarno sequence, which is conserved in bacteria. Strikingly, this entire region is absent from 12S mt-rRNA. We have mapped the ERAL1-binding site to a 33 nucleotide section delineating the 3′ terminal stem–loop region of 12S mt-rRNA. This loop contains two adenine residues that are reported to be dimethylated on mitoribosome maturation. Furthermore, and also in contrast with the bacterial orthologue, loss of ERAL1 leads to rapid decay of nascent 12S mt-rRNA, consistent with a role as a mitochondrial RNA chaperone. Finally, whereas depletion of ERAL1 leads to apoptosis, cell death occurs prior to any appreciable loss of mitochondrial protein synthesis or reduction in the stability of mitochondrial mRNA.

scholarly journals Molecular Identification of Bacterial Pathogen Infecting Coconut Leaf Beetle

Buletin Palma ◽  
2016 ◽  
Vol 16 (2) ◽  
pp. 147
Author(s):  
JELFINA C. ALOUW ◽  
DIANA NOVIANTI ◽  
MELDY L.A. HOSANG
Keyword(s):  
Polymerase Chain Reaction ◽  
16S Rrna ◽  
Serratia Marcescens ◽  
Molecular Identification ◽  
Ribosomal Rna ◽  
Causal Agent ◽  
16S Ribosomal Rna ◽  
Font Size ◽  
Chain Reaction ◽  
Polymerase Chain

<p><span style="font-size: medium;">ABSTRACT </span></p><p>Many species of microorganisms can cause diseases and mortality of insect pests. Accurate detection and identification of the entomophatogens are essential for development of biological control agent to the pest. Brontispa longissima, a serious and invasive pest of coconut, was infected by bacterium causing mortality of the larvae and pupae in coconut field. Objective of the research was to identify bacterium as a causal agent of the field-infected B. longissima using molecular  technique.  Research  was  conducted  between  April  and  August 2011.  Molecular  identification  using polymerase chain reaction (PCR) amplification of 16s ribosomal RNA of the infected larvae and sequencing of the gene showed that Serratia marcescens is the causal agent of the disease.</p><p>Keywords: Brontispa longissima, coconut, 16s rRNA, Serratia marcescens.</p><p> </p><p><span style="font-size: medium;">Identifikasi Molekular Bakteri Pathogen yang Menginfeksi Hama Daun Kelapa <br />Brontispa longissima(Coleoptera:Chrysomelidae)</span></p><p><span style="font-size: medium;">ABSTRAK </span></p><p>Banyak mikroorganisme dapat menimbulkan penyakit pada serangga hama.  Deteksi dan identifikasi yang akurat dari  pathogen  penyebab  penyakit  pada  serangga (entomopathogen)  hama  merupakan  tahap  yang  penting  dalam  pengembangan pengendalian biologi untuk hama tersebut.  Brontispa longissima sebagai hama penting dan bersifat  invasif pada tanaman kelapa diinfeksi oleh sejenis bakteri yang menyebabkan kematian larva dan pupa dari serangga  tersebut di lapangan. Penelitian ini bertujuan untuk mengidentifikasi organisme penyebab penyakit pada hama B. longissima dengan menggunakan teknik molekuler. Penelitian dilaksanakan pada bulan April sampai dengan Agustus  2011. Identifikasi bakteri dilakukan dengan mengamplifikasi 16s ribosomal RNA dari larva yang terinfeksi dengan menggunakan PCR (polymerase chain reaction).  Hasil analisis sekuens nukleotida 16s ribosomal RNA dari larva yang terinfeksi menunjukkan bahwa Serratia marcescens adalah bakteri penyebab dari penyakit tersebut.</p><p>Kata kunci: Brontispa longissima, kelapa, 16s rRNA, Serratia marcescens.</p>

Movement of the Decoding Region of the 16S Ribosomal RNA Accompanies tRNA Translocation

2000 ◽  
Vol 304 (4) ◽  
pp. 507-515 ◽  
Author(s):  
Margaret S. VanLoock ◽  
Rajendra K. Agrawal ◽  
Irene S. Gabashvili ◽  
Li Qi ◽  
Joachim Frank ◽  
...  
Keyword(s):  
Ribosomal Rna ◽  
16S Ribosomal Rna ◽  
Trna Translocation ◽  
Decoding Region

scholarly journals Molecular Diagnosis of Periprosthetic Joint Infection by Quantitative RT-PCR of Bacterial 16S Ribosomal RNA

2013 ◽  
Vol 2013 ◽  
pp. 1-4 ◽  
Author(s):  
Mel S. Lee ◽  
Wen-Hsin Chang ◽  
Su-Chin Chen ◽  
Pang-Hsin Hsieh ◽  
Hsin-Nung Shih ◽  
...  
Keyword(s):  
16S Rrna ◽  
Molecular Diagnosis ◽  
Ribosomal Rna ◽  
Periprosthetic Joint Infection ◽  
Joint Infection ◽  
16S Ribosomal Rna ◽  
Purulent Discharge ◽  
Fistula Tract ◽  
Culture Negative

The diagnosis of periprosthetic joint infection is sometimes straightforward with purulent discharge from the fistula tract communicating to the joint prosthesis. However it is often difficult to differentiate septic from aseptic loosening of prosthesis because of the high culture-negative rates in conventional microbiologic culture. This study used quantitative reverse transcription polymerase chain reaction (RT-qPCR) to amplify bacterial 16S ribosomal RNA in vitro and in 11 clinical samples. The in vitro analysis demonstrated that the RT-qPCR method was highly sensitive with the detection limit of bacterial 16S rRNA being 0.148 pg/μl. Clinical specimens were analyzed using the same protocol. The RT-qPCR was positive for bacterial detection in 8 culture-positive cases (including aerobic, anaerobic, and mycobacteria) and 2 culture-negative cases. It was negative in one case that the final diagnosis was confirmed without infection. The molecular diagnosis of bacterial infection using RT-qPCR to detect bacterial 16S rRNA around a prosthesis correlated well with the clinical findings. Based on the promising clinical results, we were attempting to differentiate bacterial species or drug-resistant strains by using species-specific primers and to detect the persistence of bacteria during the interim period before the second stage reimplantation in a larger scale of clinical subjects.

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