Year: 2012
Type: dissertation
Abstract: For about 50 years it has been known that guanine-rich sequence can, under appropriateconditions, adopt a distinctive, four-stranded, helical fold known as a G-quadruplex.Interest in quadruplex folds has grown in recent years as evidence of their biologicalrelevance has accumulated from both sequence analysis and function-specific assays.The folds are unusually stable and their formation appears to require close managementto maintain cell health; regulatory failure correlates with genomic instability and anumber of cancer phenotypes.This thesis examines, by x-ray crystallography, the solvent structure of a pre-viously reported tetramolecular RNA quadruplex, UGGGGU stabilized by Sr 2+ ions.Crystal forms of the octameric assembly formed by this sequence exhibit unusuallystrong diffraction and anomalous signal enabling the construction of reliable models toa resolution of 0.85A. The solvent structure confirms hydration patterns reported forother nucleic acid helical conformations and provides support for the greater stabilityof RNA quadruplexes relative to DNA. Novel features detected in the octameric RNAassembly include a new crystal form and evidence of multiple conformations, amongwhich one leading to the formation of a well-hydrated internal cavity.Though solvent is generally acknowledged to play a fundamental role in nucleicacid structure, its characterization from diffraction data remains challenging. To assistwith this task, the thesis investigates two novel additions to the crystallographic meth-ods arsenal. The first is segmentation of electron density maps into Morse-Smale basinscharacterized by uniform gradient flow. The second is the use of level set Fast Marchingmethods to compute the full distance field defined by the molecular surface. Both tech-niques show promise, though additional work will be required to yield effective tools.Gradient-flow segmentation provides an unambiguous way to gather all map density as-sociated with a modeled atom and enables the calculation of novel volumetric propertiesincluding total basin density. Distance field calculation provides a unified framework forcombining molecular surface calculation with surface-related queries including pocketand cavity detection and solvent travel depth.
Author: Alastair Charles Fyfe
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W1860379760