Insight into Tetramolecular DNA G-Quadruplexes Associated with ALS and FTLD: Cation Interactions and Formation of...

The G₄C₂ hexanucleotide repeat expansion in the c9orf72 gene is a major genetic cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD), with the formation of G-quadruplexes directly linked to the development of these diseases. Cations play a crucial role in the formation and structure of G-quadruplexes. In this study, we investigated the impact of biologically relevant potassium ions on G-quadruplex structures and utilized ¹⁵N-labeled ammonium cations as a substitute for K⁺ ions to gain further insights into cation binding and exchange dynamics. Through nuclear magnetic resonance spectroscopy and molecular dynamics simulations, we demonstrate that the single d(G₄C₂) repeat, in the presence of ¹⁵NH₄⁺ ions, adopts a tetramolecular G-quadruplex with an all-syn quartet at the 5'-end. The movement of ¹⁵NH₄⁺ ions through the central channel of the G-quadruplex, as well as to the bulk solution, is governed by the vacant cation binding site, in addition to the all-syn quartet at the 5'-end. Furthermore, the addition of K⁺ ions to G-quadruplexes folded in the presence of ¹⁵NH₄⁺ ions induces stacking of G-quadruplexes via their 5'-end G-quartets, leading to the formation of stable higher-ordered species.