Browsing by Author "Chilton B"

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    Controlling Topology of a Telomeric G-quadruplex DNA With a Chemical Cross-link
    (Wiley-VCH GmbH, Germany, 2025-06-18) Chilton B; Edwards PJB; Jameson GB; Hale TK; Filichev VV
    DNA G-quadruplexes (G4s) are noncanonical structures formed in guanine-rich sequences. Within the human genome, they are nonrandomly distributed and influence DNA replication, gene expression, and genome maintenance. Numerous proteins involved in these processes have been identified as G4-binding proteins. However, the interaction of proteins with G4s in the context of double-stranded DNA in vitro has been difficult to study due to the transient nature of G4s in the presence of complementary DNA. To overcome this challenge, introducing internal covalent cross-links between distant nucleotides within the DNA sequence may promote pre-folding of G4 structures, thereby shifting the thermodynamic equilibrium toward G4-formation. We used a Cu(I)-catalyzed azide–alkyne cycloaddition to create a cross-link between 2′-O-propargylguanosine and N6-azidoethyl-2′-deoxyadenosine in the DNA telomeric sequence (TAG3T)2. A cross-link between G3 and A8 reinforced the parallel G4 topology that was stable in the presence of complementary DNA. Moreover, even in the presence of its complementary strand, this cross-linked G4 recruited the parent native DNA (TAG3T)2 to form a hybrid G4. These results suggest that cross-linking provides a useful tool for stabilizing noncanonical DNA structures in the presence of complementary strands, enabling their study within the context of genomic DNA.
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    Inverted strand polarity yields thermodynamically stable G-quadruplexes and prevents duplex formation within extended DNA.
    (The Royal Society of Chemistry, 2024-08-27) Chilton B; Roach RJ; Edwards PJB; Jameson GB; Hale TK; Filichev VV
    DNA G-quadruplexes (G4) formed in guanine-rich sequences play a key role in genome function and maintenance, interacting with multiple proteins. However, structural and functional studies of G4s within duplex DNA have been challenging because of the transient nature of G4s and thermodynamic preference of G-rich DNA to form duplexes with their complementary strand rather than G4s. To overcome these challenges, we have incorporated native nucleotides in G-rich sequences using commercially available inverted 3'-O-DMT-5'-O-phosphoramidites of native nucleosides, to give 3'-3' and 5'-5' linkages in the centre of the G-tract. Using circular dichroism and 1H nuclear magnetic resonance spectroscopies and native gel electrophoresis, we demonstrate that these polarity-inverted DNA sequences containing four telomeric repeats form G4s of parallel topology with one lateral or diagonal loop across the face of the quadruplex and two propeller loops across the edges of the quadruplex. These G4s were stable even in the presence of complementary C-rich DNA. As an example, G4 assemblies of inverted polarity were shown to bind to the hinge region of Heterochromatin Protein 1α (HP1α), a known G4-interacting domain. As such, internal polarity inversions in DNA provide a useful tool to control G4 topology while also disrupting the formation of other secondary structures, particularly the canonical duplex.