Multi-scale Simulation Of Folding Pathway Of Adenine Riboswitch

Riboswitches are noncoding mRNA elements that can regulate the gene expression via altering their structures in response to specific metabolite binding. Therefore, elucidating and characterizing the folding/unfolding process of riboswitches are very important for understanding their regulatory mechanisms.In this dissertation we studied the unfolding behavior of add A-riboswitch through the iterative topology-based Gaussian network model. In this model the unfolding behavior of add A-riboswitch was mimicked by breaking the native contacts between different nucleotides according to the fluctuations of the distance between them. The complete sequence of add A-riboswitch unfolding event was obtained, which is consistent well with the experimental data of single molecule force measurements. Since only the native contacts are considered during the simulation, this result suggests that the native topology may play an important role in the add A- riboswitch unfolding process.Additionally, steered molecular dynamics simulation was used to explore the unfolding pathway of add A-riboswitch, too. Molecular dynamics simulation is an important method to investigate the detailed microscopic dynamics of biomolecules and has been widely used in the field of dynamic studies of biological macromolecules. However, due to the expensive computational costs, it is hard to simulate the whole unfolding process. Instead we used steered molecular dynamics simulation to explore the unfolding behavior, which was speeded up largely by external force; therefore the simulation can be done in acceptable time scale.There are two contents in this thesis:(1) Explore the unfolding pathway of add A-riboswitch with the iterative Gaussian network model. Using the iterative Gaussian network model proposed by our group in studying protein unfolding pathway, we simulated the add A-riboswitch’s unfolding behavior and analyzed metal ion and ligand’s effects onthe native conformation. Instead of using P atoms as nodes as in conventional Gaussian network model method, we chose the atomsof bases to mark different types of nucleotides. These mark atoms was chosen based on Watson-Crick base-pairing. The results of iterative simulations show a complete unfolding pathway which is consistent with the experimental data. After that we analyzed the change of cross-correlations between nucleotide fluctuations during the unfolding process, and the results indicate the denatured state seems to move highly cooperatively. In addition, we investigated fluctuations’ change in the fast modes during unfolding process. It’s found that regions close to binding pocket hold higher fluctuations. At last we analyzed the effects of Mg2+ ions and the ligand on the dynamical properties of add A-riboswitch. The results display that they are helpful in pre-organization in some degree, as well as helpful in stabilizing the folded conformation of the riboswitch.(2) Study the unfolding process of add A-riboswitch by steered molecular dynamics simulations. In order to obtain the complete unfolding process in acceptable time scale,the external force was put on the riboswitch molecular. The unfolding pathway we got with steered molecular dynamics simulation is roughly consistent with experimental results. In additional we analyzed the changes of interactions between atoms in loop-loop region and junctions region. The result indicate that the base stacking interactions is reduced when the ligand gone.