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16S Ribosomal RNA

The Experts below are selected from a list of 249 Experts worldwide ranked by ideXlab platform

Jean-pierre Goudonnet – 1st expert on this subject based on the ideXlab platform

  • Scanning tunnelling microscopy of 16S Ribosomal RNA in water.
    Biochemical and Biophysical Research Communications, 1991
    Co-Authors: Eric Lesniewska, Pierre-jacques Flamion, Claire Cachia, Jean-pierre Schreiber, Jean-pierre Goudonnet

    Abstract:

    Abstract The scanning tunnelling microscope has been used to image 16S Ribosomal RNA molecules in water electrophoretically deposited on graphite surface. Two kinds of images have been obtained: images showing aggregates of 16S Ribosomal RNA molecules similar to those obtained from DNA solutions and others showing individual 16S Ribosomal RNA molecules. An interesting characteristic of these images, recorded in constant current mode, is that the 16S Ribosomal RNA molecules appear to be located below the graphite surface. The morphology and several structural parameters of the molecules were consistent with the data obtained from electron microscopy.

  • Images of 16S Ribosomal RNA by scanning tunnelling microscopy.
    Journal of microscopy, 1991
    Co-Authors: Pierre-jacques Flamion, Eric Lesniewska, Claire Cachia, Jean-pierre Schreiber, T David, Jean-pierre Goudonnet

    Abstract:

    We report the use of scanning tunnelling microscopy (STM) to study surface topographies of complex nucleic acid structures. From low-resolution STM images of uncoated 16S Ribosomal RNA, we demonstrate the possibility of determining several objective parameters (molecular mass and radius of gyration) in order to characterize and identify the molecules observed. These parameters were compared with values obtained by other physical methods and the radius of gyration was found to be the most reliable. At high resolution, it was possible to measure the main dimensions of selected V-form particles more precisely than with electron microscopy. Images of the more compact form have been also obtained that show different domains in the macromolecular structure.

Harry F Noller – 2nd expert on this subject based on the ideXlab platform

  • Directed hydroxyl radical probing of 16S Ribosomal RNA in 70S ribosomes from inteRNAl positions of the RNA.
    Biochemistry, 1999
    Co-Authors: Lisa F. Newcomb, Harry F Noller

    Abstract:

    : Directed hydroxyl radical probing of 16S Ribosomal RNA from Fe(II) tethered to specific sites within the RNA was used to determine RNARNA proximities in 70S ribosomes. We have transcribed 16S Ribosomal RNA in vitro as two separate fragments, covalently attached an Fe(II) probe to a 5′-guanosine-alpha-phosphorothioate at the junction between the two fragments, and reconstituted 30S subunits with the two separate pieces of RNA and the small subunit proteins. Reconstituted 30S subunits capable of association with 50S subunits were selected by isolation of 70S ribosomes. Hydroxyl radicals, generated in situ from the tethered Fe(II), cleaved sites in the 16S rRNA backbone that were close in three-dimensional space to the Fe(II), and a primer extension was used to identify these sites of cleavage. Two sets of 16S Ribosomal RNA fragments, 1-360/361-1542 and 1-448/449-1542, were reconstituted into active 30S subunits. Fe(II) tethered to position 361 results in cleavage of 16S rRNA around nucleotides 34, 160, 497, 512, 520, 537, 552, and 615, as well as around positions 1410, 1422, 1480, and 1490. Fe(II) tethered to position 449 induces cleavage around nucleotide 488 and around positions 42 and 617. Fe(II) tethered to the 5′ end of 16S rRNA induces cleavage of the rRNA around nucleotides 5, 601, 615, and 642. These results provide constraints for the positioning of these regions of 16S rRNA, for which there has previously been only limited structural information, within the 30S subunit.

  • Directed hydroxyl radical probing of 16S Ribosomal RNA in ribosomes containing Fe(II) tethered to Ribosomal protein S20.
    RNA, 1998
    Co-Authors: Gloria M. Culver, Harry F Noller

    Abstract:

    The 16S Ribosomal RNA neighborhood of Ribosomal protein S20 has been mapped, in both 30S subunits and 70S ribosomes, using directed hydroxyl radical probing. Cysteine residues were introduced at amino acid positions 14, 23, 49, and 57 of S20, and used for tethering 1-(p-bromoacetamidobenzyl)-Fe(II)-EDTA. In vitro reconstitution using Fe(II)-derivatized S20, together with the remaining small subunit Ribosomal proteins and 16S Ribosomal RNA (rRNA), yielded functional 30S subunits. Both 30S subunits and 70S ribosomes containing Fe(II)-S20 were purified and hydroxyl radicals were generated from the tethered Fe(II). Hydroxyl radical cleavage of the 16S rRNA backbone was monitored by primer extension. Different cleavage patterns in 16S rRNA were observed from Fe(II) tethered to each of the four positions, and these patterns were not significantly different in 30S and 70S ribosomes. Cleavage sites were mapped to positions 160-200, 320, and 340-350 in the 5′ domain, and to positions 1427-1430 and 1439-1458 in the distal end of the penultimate stem of 16S rRNA, placing these regions near each other in three dimensions. These results are consistent with previous footprinting data that localized S20 near these 16S rRNA elements, providing evidence that S20, like S17, is located near the bottom of the 30S subunit.

  • a functional pseudoknot in 16S Ribosomal RNA
    The EMBO Journal, 1991
    Co-Authors: Ted Powers, Harry F Noller

    Abstract:

    Several lines of evidence indicate that the universally conserved 530 loop of 16S Ribosomal RNA plays a crucial role in translation, related to the binding of tRNA to the Ribosomal A site. Based upon limited phylogenetic sequence variation, Woese and Gutell (1989) have proposed that residues 524-526 in the 530 hairpin loop are base paired with residues 505-507 in an adjoining bulge loop, suggesting that this region of 16S rRNA folds into a pseudoknot structure. Here, we demonstrate that Watson-Crick interactions between these nucleotides are essential for Ribosomal function. Moreover, we find that certain mild perturbations of the structure, for example, creation of G-U wobble pairs, generate resistance to streptomycin, an antibiotic known to interfere with the decoding process. Chemical probing of mutant ribosomes from streptomycin-resistant cells shows that the mutant ribosomes have a reduced affinity for streptomycin, even though streptomycin is thought to interact with a site on the 30S subunit that is distinct from the 530 region. Data from earlier in vitro assembly studies suggest that the pseudoknot structure is stabilized by Ribosomal protein S12, mutations in which have long been known to confer streptomycin resistance and dependence.

Heidi H Kong – 3rd expert on this subject based on the ideXlab platform

  • Research Techniques Made Simple: Bacterial 16S Ribosomal RNA Gene Sequencing in Cutaneous Research
    Journal of Investigative Dermatology, 2015
    Co-Authors: Jay-Hyun Jo, E A Kennedy, Heidi H Kong

    Abstract:

    Skin serves as a protective barrier and also harbors numerous microorganisms collectively comprising the skin microbiome. As a result of recent advances in sequencing (next-generation sequencing), our understanding of microbial communities on skin has advanced substantially. In particular, the 16S Ribosomal RNA gene sequencing technique has played an important role in efforts to identify the global communities of bacteria in healthy individuals and patients with various disorders in multiple topographical regions over the skin surface. Here, we describe basic principles, study design, and a workflow of 16S Ribosomal RNA gene sequencing methodology, primarily for investigators who are not familiar with this approach. This article will also discuss some applications and challenges of 16S Ribosomal RNA sequencing as well as directions for future development.