The influence of G-tract and loop length on the topological variability of putative five and six G-quartet DNA structures in the human genome

Local variation of DNA structure and its dynamic nature play an essential role in the regulation of important biological processes. One of the most prominent noncanonical structures are G-quadruplexes, which form in vivo within guanine-rich regions and have been demonstrated to be involved in the regulation of transcription, translation and telomere maintenance. We provide an analysis of G-quadruplex formation in sequences with five and six guanine residues long G-tracts, which have emerged from the investigation of the gapless human genome and are associated with genes related to cancer and neurodegenerative diseases. We systematically explored the effect of G-tract and loop elongations by means of NMR and CD spectroscopy and polyacrylamide electrophoresis. Despite both types of elongation leading up to structural polymorphism, we successfully determined the topologies of four out of eight examined sequences, one of which contributes to a very scarce selection of currently known intramolecular four G-quartet structures in potassium solutions. We demonstrate that examined sequences are incompatible with five or six G-quartet structures with propeller loops, although the compatibility with other loop types cannot be factored out. Lastly, we propose a novel approach towards specific G-quadruplex targeting that could be implemented in structures with more than four G-quartets. •Putative five and six G-quartet sequences are rare and appear in T2T genome.•Five and six G-quartet structures are incompatible with propeller loops.•T(G4T)4 adopts a parallel three-quartet structure.•T(G5T)4 adopts an antiparallel four-quartet structure.•Extending G-tract above five residues promotes intermolecular species.