Abstract
Ribosome biogenesis in E. coli has been extensively studied, yet, very little is known about the structural changes that occur to the rRNA upon r-protein binding. Recent cryo-electron microscopy and mass spectrometry studies from the Williamson laboratory have produced a time-resolved assembly map of the 16S subunit. The first r-protein to nucleate 16S folding, S4, binds to h16-h18 of the five-way junction–a signature region that idiosyncratically separates Bacteria and Archaea. S4, subsequently, nucleates the formation of the 5' domain of the 16S and recruits other r-proteins to bind.Guided by in vitro studies, we probe how S4 nucleates the 5' domain formation using a series of all-atom molecular dynamics (MD) simulations on the E. coli 5' domain with various r-proteins systematically bound to the rRNA. Large dynamic fluctuations in the structure of the rRNA were observed in both our long-time MD trajectories and multi-color single molecule FRET experiments from our collaborators. For example, with only S4 bound to the rRNA, an A-minor interaction between h8 and h14 opens up, but is stable upon adding S17 to a location distant from h8 and h14.We also are exploring the cooperative effects of the r-protein binding by generating weighted networks based on position correlations. Graph of the network weighed by edge-betweenness, a measure of the number of shortest paths that pass through a given edge, show a continuous path connecting the binding sites of S4, S17, and S20 with a branching point located near h15–the binding site the secondary binding r-protein S16. This path is hypothesized to modulate the binding between primary and secondary binding r-proteins and further probed using additional all-atom and structure-based models.
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