Structural characterisation of nucleic acid quadruplexes

Abstract

Guanine-rich sequences of DNA or RNA have the potential to form higher order structural motifs known as G-quadruplexes (G4s). They have been implicated in a variety of biological processes, adding an exquisite level of refinement to the control of gene expression and have also shown utility in the fields of nanotherapeutics and nanotechnology. Canonical G-quadruplexes are comprised of four tracts of equal amounts of guanosines, with all guanines engaged in a stem of stacked G-tetrads, linked by three loops. They can be highly sensitive to changes in their solution environment or to small mutations in the sequence of their loops. Therefore, polymorphism is pronounced not only for G4s formed by different sequences but may also be observed for a given sequence. G-quadruplex polymorphism enhances functionality but frustrates their synthetic assembly. As such, there is a pressing need to unravel the inherent dynamic and polymorphic nature of G4 motifs and to develop a comprehensive set of rules for their prediction and thus fabrication. The work described herein is primarily concerned with the evaluation of the topological characteristics of a unimolecular, and a tetrameric G-quadruplex architectures of DNA, as well as an RNA tetrameric ‘kissing complex’. The topological analysis is derived from resonance assignments of nuclei in multidimentional, multinuclear NMR experiments.

 The RNA tetrameric complex is the first architecture of RNA in which G-quadruplexes recognize each other simply through the stacking of tetrads- a ‘kissing complex’. In the tetrameric DNA topology the first (C:G:G:G:G:C) hexad architecture is observed. This feature is observed in a quadruplex stem that involves symmetry-related strands linked by (G:C:G:C) tetrads. The unimolecular G-quadruplex architecture is only the second architecture determined to atomic detail, that having the potential to form an N- number of tetrads, is only able to form N-1 tetrads (collapse). This specific collapse, the formation of the RNA ‘kissing complex’, and formation of the novel hexad are here discussed in the context of control of self-assembly. There is thus ongoing work on assessing self-assembly by other methods.

Thesis embargoed until 31 May 2027

Date of Award	May 2025
Original language	English
Supervisor	Steven Watterson (Supervisor) & Mateus Webba Da Silva (Supervisor)