Date of Award




Document Type

Master's Thesis

Degree Name

Master of Science (MS)


Department of Chemistry

Content Description

1 online resource (x, 119 pages) : color illustrations.

Dissertation/Thesis Chair

Alan A.C. Chen

Committee Members

Alexander A.S. Shekhtman, Jia J.S. Sheng, Mehmet M.Y. Yigit


MicroRNA, Messenger RNA, Proteins, Nuclear magnetic resonance, Protein folding

Subject Categories

Biophysics | Computational Chemistry


MicroRNA (miRNA), as a distinct class of biological regulators and a ”guide” member of non-coding RNA-protein complexes (RNPs), regulates more than 60% of protein-coding genes expression through base-pairing with targeted messenger RNA (mRNA) in the RNA-Induced Silencing Complex (RISC). Most of miRNAs identified in human, are conserved in other animals, which have preferentially conserved interaction sites particularly in 3’ untranslated regions (3’UTRs) of many human messenger mRNAs.The capability of a single miRNA to target more than hundreds of mRNAs, suggests that miRNAs influence essentially all developmental process and diseases, which also makes them interesting candidates as therapeutics agents. The primary determinant of miRNA recognition is the level of sequence complementarity in the corresponding binding site of mRNA. However, the precise mechanism in which RISC is activated and repress the translation of mRNA remained a mystery using available experimental techniques. We hypothesize that the miRNA-mRNA interaction does not occur only at the level of sequence complementarity. Moreover, it may profit from inherent structural flexibility of imperfect miRNA-mRNA pairing, which encodes for functionally relevant conformational rearrangements of this pair, directing RISC activation. Among experimental tools, nuclear magnetic resonance (NMR) measurements are able to collect conformational dynamics of biomolecules on various timescales. Such measurements, for RNAs with highly flexible and multi-state nature, particularly in systems with the size and heterogeneity of RISC, are precluded with local, incomplete structural dynamics at the secondary level. Molecular Dynamics (MD) simulations are able to complement these experimental measurements, by contextualizing how the 3D architecture and atomic motions of RNAs give rise to their molecular function. The accuracy of simulation results are mainly relies on the employed energy function and fully converged sampling. Despite of steadily improvements and developments of force fields and computation power, simulating the RNA conformation space remains challenging. Technically, assessing the accuracy of a force field, mainly depends on the consistency of simulation results with the available experimental data. In this work, primarily we assess the accuracy of selected RNA force fields with varied strength base-pairs, against thermodynamics of the site-specific Sallmonella 4U thermometer. Secondarily, we present a method to incorporate sparse base-pairing information to dramatically improve the accuracy and efficiency of 3D RNA folding using a novel 2D replica exchange protocol. We will show that just by incorporating a handful of secondary structure restraints into an otherwise unbiased all-atom simulation, accurate RNA tertiary folds can be modelled at the fraction of the effort of de-novo simulation of RNA folding. This method can potentially incorporate base-pairing information from a wide variety of experimental inputs to predict highly complex RNA tertiary folds including pseudoknots, multi-loop junctions, and non-canonical interactions. Then, using the simulation groundwork we have developed and introduced earlier, we show how the 3D structural dynamics of the NMR observed conformational transition of miRNA–mRNA targeting persuade further conformational change by Argonaute2 (Ago2) required for RISC function. Lastly, we use molecular dynamics simulations to study the conformation dynamics of DNA/RNA assemblies consisting of varied number of 2’-5’ linkage modification in the RNA components of these nanostructures. The naturally occurring 2’-5’ modification with certain features such as nuclease resistance and binding specificity, while maintaining stability, canadvance the functionality of nanostructures in medicine and drug delivery, especially those requiring strand release.