Bao, ChenLoerch, SarahLing, ClarenceKorostelev, Andrei A.Grigorieff, NikolausErmolenko, Dmitri N.2022-08-232022-08-232020-02-062020-03-23<p>bioRxiv 2020.02.05.936120; doi: https://doi.org/10.1101/2020.02.05.936120. <a href="https://doi.org/10.1101/2020.02.05.936120" target="_blank">Link to preprint on bioRxiv service.</a></p>10.1101/2020.02.05.936120https://hdl.handle.net/20.500.14038/29445Although the elongating ribosome is an efficient helicase, certain mRNA stem-loop structures are known to impede ribosome movement along mRNA and stimulate programmed ribosome frameshifting via mechanisms that are not well understood. Using biochemical and single-molecule Förster resonance energy transfer (smFRET) experiments, we studied how frameshift-inducing stem-loops from E. coli dnaX mRNA and the gag-pol transcript of Human Immunodeficiency Virus (HIV) perturb translation elongation. We find that upon encountering the ribosome, the stem-loops strongly inhibit A-site tRNA binding and ribosome intersubunit rotation that accompanies translation elongation. Electron cryo-microscopy (cryo-EM) reveals that the HIV stem-loop docks into the A site of the ribosome. Our results suggest that mRNA stem-loops can transiently escape ribosome helicase by binding to the A site. Thus, the stem-loops can modulate gene expression by sterically hindering tRNA binding and inhibiting translation elongation.en-USThe copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It is made available under a CC-BY-ND 4.0 International license.http://creativecommons.org/licenses/by-nd/4.0/BiochemistrymRNAribosomestRNA bindingframeshift-inducing stem-loopshelicasesFörster resonance energy transferE. coli dnaXHuman Immunodeficiency VirusBiochemical Phenomena, Metabolism, and NutritionBiochemistryEnzymes and CoenzymesGenetic PhenomenaNucleic Acids, Nucleotides, and NucleosidesmRNA stem-loops can pause the ribosome by hindering A-site tRNA binding [preprint]Preprinthttps://escholarship.umassmed.edu/cgi/viewcontent.cgi?article=2679&context=faculty_pubs&unstamped=1https://escholarship.umassmed.edu/faculty_pubs/167016954038faculty_pubs/1670