Towards automating structural analysis of complex RNA molecules and some applications in nanotechnology

The unifying theme of this dissertation is to develop new conceptual frameworks and analytical approaches to assess and improve the automated annotation and analysis of RNA 3D structures, and to connect these data to structural changes in functional state and evolutionary changes. More specifically, the dissertation focuses on 3D structures of ribosomal RNA (rRNA) from the large (LSU) and small (SSU) subunits of ribosomes, and the hairpin and internal loop motifs extracted from them. As a first step toward automated annotation of the functional states of ribosome structures, a manual analysis was carried out of all atomic-resolution, 3D structures of the SSU of the bacterium T. thermophilus found in the NDB as of April, 2014. The data set used corresponds to the structure equivalence class NR_4.0_81883.24 of release 1.56 of ribosome structures posted in the NDB (see http://rna.bgsu.edu/rna3dhub/nrlist). Each structure was manually examined to determine the functional annotations for the state of the ribosome with all bound tRNAs, mRNAs and other factors. These data were combined with meta-data downloaded from NDB in a single spreadsheet found at this link: http://tinyurl.com/16S-T-Thermophilus-summary.;NDB maintains a Non-redundant (NR) set of structures that is updated each week and used to construct the 3D Motif Atlas of RNA hairpin and internal loop motifs (http://rna.bgsu.edu/rna3dhub/motifs), updated monthly. To assess the quality of motif clustering in the Motif Atlas, links and meta-data were downloaded for all motif instances in release 1.14 (http://rna.bgsu.edu/rna3dhub/motifs/release/IL/1.14). Motifs were grouped by molecule and location within each homologous molecule to determine whether homologous motifs were clustered in the Motif Atlas in the same or similar structural groups (identified by motif ID). The focus of analysis was rRNA for which multiple instances from different organisms are available. Overall, motifs found in conserved regions of the rRNAs were placed in the same motif groups by the automated clustering.;Large RNA structures like the ribosome are organized hierarchically into subunits, domains, stacked helical units and individual helices and their component hairpin and internal loop motifs. Different parts of the structure are responsible for binding mRNA, tRNA and translation factors, but all must work together to correctly translate the mRNA. For example, the amino-acyl end of a tRNA should not be allowed into the A-site of the 23S rRNA unless the 16S rRNA detects cognate binding between the anti-codon end of the tRNA and the mRNA codon present in the decoding site, detection done in helix 44. Thus, information must flow between different parts of the ribosome to report on the state of one part that is relevant to another part. How this happens in complex RNA machines is an area of active research. One hypothesis is that networks of RNA tertiary and quaternary interactions play a central role in the ribosome. As a first step to automatically detecting and comparing networks of RNA-RNA interactions in the ribosome, a new module was created for the FR3D ("Find RNA 3D") suite of RNA analysis tools. The module takes as input CSV formatted data, listing the manually constructed, hierarchical assignments of each nt in an RNA 3D structure, and the atomic-resolution PDB structure of the molecule. The program outputs a table with the number of interactions between elements, individual PDB files for each pair of interacting elements for visual inspection, and a text file with the list of interactions between elements. Thus, it is possible to analyze large structures automatically. (Abstract shortened by UMI.).