CONTINUOUS LONG-TIME DYNAMICS OF RNA MOLECULES: WATCHING WITHOUT BLINK- ING FOR

Carnegie-Mellon University, Pittsburgh PA

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Here we propose to generate a continuous 100 ?s trajectory of HIV-1 TAR, a prototypical non-coding RNA system. Importantly, we will make direct connection to liquid state NMR measurements that have important sensitivity to structural dynamics rather than static disorder. While the latter could be generated with trivial parallel computing, we need to be able to simulate the correct, continuous dynamics. Two main issues will be addressed: 1) Residual dipolar coupling, which probes picosecond to millisecond dynamics, will be calculated and compared to experiment from the lab of our collaborator, Prof Al-Hashimi (Univ. of Michigan). Comparisons where the RDCs were calculated from trajectories on the order of nanoseconds exhibit poor agreement with experimentally determined RDCs. This observation behooves one to ask whether whether the poor agreement was due to the fact that on the nanosecond scale the systems were non-ergodic and thus conformational space was not sampled with the correct distribution, or it was due to the inaccuracies of the underlying forcefield that generated incorrect dynamics (or both). 2) TAR contains two non-canonical RNA motifs, an internal bulge and an apical loop. Experimental evidence from Al-Hashimi group suggest that the bulge and the loop undergo dynamics on the microsecond timescale. It is believe that these motions involve inter-conversion between various metastable states. For both the bulge and loop, conformational space network analysis will be used to explore the free energy landscape so as to identify these metastable states, the exact sequencing of intermediates, and the rates of conversion between them. These questions, for which we seek answers, can only be addressed by the generation and then analysis of continuous trajectories on the order of hundreds on microseconds. Moreover, the actual dynamics can be used to discern between the various motional models that are proposed for RNA structural changes. Generation of the required trajectory with standard supercomputers will require substantial computational resources and CPU time, and with standard queuing practices, will take many months to complete. Use of Anton at NRBSC will allow us to generate the required trajectory in less than a week.