Theses and Dissertations from UMD

Permanent URI for this communityhttp://hdl.handle.net/1903/2

New submissions to the thesis/dissertation collections are added automatically as they are received from the Graduate School. Currently, the Graduate School deposits all theses and dissertations from a given semester after the official graduation date. This means that there may be up to a 4 month delay in the appearance of a give thesis/dissertation in DRUM

More information is available at Theses and Dissertations at University of Maryland Libraries.

Browse

Filters

2006 - 200912010 - 20151

Settings

Subject: search.filters.subject.translational fidelity

Search Results

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    The Core of Eukaryotic Ribosomal Protein uS19 Functions as a Pivot Point Enhancing Eukaryotic Ribosome Flexibility
    (2015) Bowen, Alicia Marie; Dinman, Jonathan D; Biochemistry; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    While most ribosomal elements are highly conserved in the three domains of life, over the course of evolution, significant differences have emerged as ribosomes have been subjected to different types of selective pressure. In prokaryotes and archaea, a single small subunit protein, uS13, partners with H38 (the A-site finger) and uL5 to form the B1a and B1b/c bridges, respectively. In eukaryotes, it appears that the small subunit component was split into two separate proteins during the course of evolution. One of these, also known as uS13 (previously known as S18), only participates in bridge B1b/c with uL5 in eukaryotes (previously known as L11). The other, called uS19 (previously known as S15) is the small subunit partner in the B1a bridge with H38. Here, poly-alanine mutants of the uS19/Us13 interface of uS19 were used to elucidate the evolutionary advantage of this split. A previously described chemical protection profile of the B7a bridge was utilized in order to determine the ribosomal rotational status of the selected mutants. rRNA structure probing analyses reveal that uS19/uS13 interface mutations shift the ribosomal rotational equilibrium toward the unrotated state. This perturbation of ribosomal rotational equilibrium also affected the ribosomes affinity for two intrinsic ligands: unrotated ribosomes exhibit increased affinity for ternary complex and disfavor binding of the translocase, eEF2. We posit that this mutation causes the normally flexible head region of the small subunit to "stiffen", thereby decreasing the ribosomes range of motion. A model is presented in which residues L112GH114 at the uS19/uS13 interface act as the ball in a "ball-and-socket joint", providing the increased flexibility required in the head region of the eukaryotic SSU as a consequence of the evolutionary process.
  • Thumbnail Image
    Item
    Ribosome integrity and translational fidelity require accurate modification and processing of rRNA in the yeast Saccharomyces cerevisiae
    (2006-11-21) Roshek, Jennifer Lynn Baxter; Dinman, Jonathan D; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Translating mRNA sequences into functional proteins is a fundamental process necessary for the viability of organisms throughout all kingdoms of life. The ribosome carries out this process with a delicate balance between speed and accuracy. Although kinetic and biochemical studies along with high resolution crystal structures have provided much information about the ribosome, many of the underlying mechanisms of ribosome function are still poorly understood. This work seeks to understand how ribosome structure and function are affected by changes in rRNA as caused by two very different mechanisms. mof6-1, originally isolated as a recessive mutation which promoted increased efficiencies of programmed -1 ribosomal frameshifting, was found to be an allele of RPD3 which encodes a histone deacetylase that is involved in transcriptional activation and silencing. This mutant demonstrated a delay in ribosomal RNA (rRNA) processing leading to changes in reading frame maintenance and ribosomal A-site specific defects. To understand the role of cis-acting changes to rRNA, yeast strains deficient in rRNA modifications in the peptidyl transferase center of the ribosome were monitored for changes in ribosome structure and translational fidelity. Analyses revealed mutant phenotypes including sensitivity to translational inhibitors; changes in reading frame maintenance, nonsense suppression and aa-tRNA selection; and increased rates of A-site tRNA binding to the mutant ribosome. One mutant in particular, spb1DA/snr52Δ, promoted increased rates of programmed -1 ribosomal frameshifting, increased rates of near cognate tRNA selection and A-site tRNA binding. Structural analysis of spb1DA/snr52Δ revealed changes consistent with a more accessible ribosomal A-site. These results suggest that rRNA nucleotide modifications produce small but distinct changes in ribosome structure and function contributing to overall translational fidelity. Taken together, these data suggest that rRNA, a main component of the ribosome, contributes directly to translational fidelity. Defects in rRNA caused by changes in both its processing and modification can cause changes in reading frame maintenance, nonsense suppression, aa-tRNA selection and binding as well as ribosome structure.