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By using single-molecule fluorescence resonance energy transfer (smFRET), we observe the real-time dynamic coupling between the ribosome, labeled at the L1 stalk, and transfer RNA (tRNA). We find that an interaction between the ribosomal L1 stalk and the newly deacylated tRNA is established spontaneously upon peptide bond formation; this event involves(More)
Time series data provided by single-molecule Förster resonance energy transfer (smFRET) experiments offer the opportunity to infer not only model parameters describing molecular complexes, e.g., rate constants, but also information about the model itself, e.g., the number of conformational states. Resolving whether such states exist or how many of them(More)
Ribosomal frameshifting occurs when a ribosome slips a few nucleotides on an mRNA and generates a new sequence of amino acids. Programmed -1 ribosomal frameshifting (-1PRF) is used in various systems to express two or more proteins from a single mRNA at precisely regulated levels. We used single-molecule fluorescence resonance energy transfer (smFRET) to(More)
Determining the mechanism by which tRNAs rapidly and precisely transit through the ribosomal A, P, and E sites during translation remains a major goal in the study of protein synthesis. Here, we report the real-time dynamics of the L1 stalk, a structural element of the large ribosomal subunit that is implicated in directing tRNA movements during(More)
Translocation of tRNAs through the ribosome during protein synthesis involves large-scale structural rearrangement of the ribosome and ribosome-bound tRNAs that is accompanied by extensive and dynamic remodeling of tRNA-ribosome interactions. How the rearrangement of individual tRNA-ribosome interactions influences tRNA movement during translocation,(More)
Cells express many ribosome-interacting factors whose functions and molecular mechanisms remain unknown. Here, we elucidate the mechanism of a newly characterized regulatory translation factor, energy-dependent translational throttle A (EttA), which is an Escherichia coli representative of the ATP-binding cassette F (ABC-F) protein family. Using cryo-EM, we(More)
The use of aptamer-fluorogen complexes is an emerging strategy for RNA imaging. Despite its promise for cellular imaging and sensing, the low fluorescence intensity of the Spinach-DFHBI RNA aptamer-fluorogen complex hampers its utility in quantitative live-cell and high-resolution imaging applications. Here we report that illumination of the(More)
Base-pairing interactions between nucleic acids mediate target recognition in many biological processes. We developed a super-resolution imaging and modeling platform that enabled the in vivo determination of base pairing-mediated target recognition kinetics. We examined a stress-induced bacterial small RNA, SgrS, which induces the degradation of target(More)
Live cell RNA imaging using genetically encoded fluorescent labels is an important tool for monitoring RNA activities. A recently reported RNA aptamer-fluorogen system, the Spinach, in which an RNA aptamer binds and induces the fluorescence of a GFP-like 3,5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI) ligand, can be readily tagged to the RNA of(More)
Characterizing the structural dynamics of the translating ribosome remains a major goal in the study of protein synthesis. Deacylation of peptidyl-tRNA during translation elongation triggers fluctuations of the pretranslocation ribosomal complex between two global conformational states. Elongation factor G-mediated control of the resulting dynamic(More)