Please use this identifier to cite or link to this item: https://hdl.handle.net/10356/73074
Title: Investigating the solution structure of G-rich DNA and RNA using AFM
Authors: Bose, Krishnashish
Keywords: DRNTU::Science::Biological sciences::Biophysics
DRNTU::Science::Physics::Atomic physics::Solid state physics
Issue Date: 2018
Source: Bose, K. (2018). Investigating the solution structure of G-rich DNA and RNA using AFM. Doctoral thesis, Nanyang Technological University, Singapore.
Abstract: G-rich DNA/RNA sequences are highly dynamic and diverse in nature as they can adopt a myriad of secondary structures in physiologically relevant conditions. Several biologically relevant G-rich repeat motif DNA sequences are known to form higher-order G-quadruplexes, but little is known about their atomic level structure. Atomic Force Microscopy (AFM) has been widely used to investigate higher-order G-quadruplexes for the past 3 decades. In the current work, AFM has been pushed to the limits to develop a new methodology for probing biomolecular structure with sub-nanometer resolution and thus bridge the gap between single-molecule and high-resolution ensemble techniques. The protocol for visualization of G-quadruplex DNA and RNA in aqueous solution was developed to resolve their intricate structural features. Higher-order structures formed by three different biologically relevant sequences containing G-rich repeat motif were investigated, viz., GGGGTT, GGGGCC and TTAGGG. It was found that short and long G-wires are formed by d[G4T2G4], d[G4C2G4] and r[G4C2G4]. All these G-wires revealed similar morphology (uniform height, variable length) and periodic surface pattern. We built several molecular models and found one that can explain the experimentally observed features of G-wires. Our work hint towards a unified architecture adopted by G-wires formed by different DNA and RNA oligonucleotides. These findings are useful for the design of G-wire based nanodevices and may find relevance in biology. Transcription of PCR-amplified DNA containing several repeats of TTAGGG was used to generate telomeric repeat containing RNA (TERRA), which was investigated using high-resolution AFM in aqueous solution. The higher-order structures formed by TERRA were grouped into two broad classes based on their heights and intricate structural periodicities. One class of observed structures were like the pearl necklace as proposed earlier for telomeric DNA and RNA. In summary, we conclude that G-rich DNA and RNA sequences are capable of self-assembling into higher-order intermolecular G-quadruplexes which are consistent with a model of stacked G-tetrads having molecular periodicities which are multiples of the distance between individual G-tetrad layers. However, individual strands of G-rich DNA and RNA could exist as a pearl necklace, where the beads are likely G-quadruplexes, which are connected by a single-stranded linker. The higher-order structures reported here could be broadly classified into just two groups: four-stranded parallel G-wire and single-stranded pearl necklace.
URI: http://hdl.handle.net/10356/73074
DOI: 10.32657/10356/73074
Fulltext Permission: open
Fulltext Availability: With Fulltext
Appears in Collections:SPMS Theses

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