Quantifying the Stability of Acridines to Ribosomal G-Quadruplexes

Abstract

G-quadruplexes are involved in fundamental regulatory processes, including those associated with cancer. Previous studies primarily focused on telomeric and oncogenic DNA G-quadruplexes, but recent studies have demonstrated that G-quadruplexes in ribosomal DNA or ribosomal RNA (rDNA or rRNA), may be an alternative target with a significant clinical potential through the inhibition of RNA polymerase I (Pol I) in ribosome biogenesis. The structures of ribosomal G-quadruplexes at atomic resolution are unknown and very little biophysical characterization has been performed on them. In our present study, we modeled putative ribosomal G-quadruplexes that were previously predicted using a bioinformatics algorithm and verified using CD spectroscopy to exist in a predominantly parallel topology. To quantify the relative stabilities of the acridine:G-quadruplex structures, we introduce two novel metrics for quantifying the relative stability of the G-quadruplex tetrads: 1) the center of mass base-to-base distance between diagonal guanines, and 2) the torsional angle between four guanines. Our relative free energy profiles show that the rDNA G-quadruplex structures with shorter loops are more stable for parallel topologies. The antiparallel topology was determined adopt a disordered configuration due to the lack of planarity after the simulation. To evaluate acridine molecules as a viable small molecule chemotherapeutic agent, we then rationally designed several acridine molecules. Each has a polyaromatic face that binds to the top-most G-tetrad and has amino or carboxyl side-chains to selectively optimize the shape and electrostatic complementarity. We then docked acridines to the rDNA G-quadruplexes and performed MD simulations. We identified several acridines with amino groups, and the nature of their specificity and relative stabilities depend on the rDNA G-quadruplex loop structures.