Interactions between a Bioflavonoid and G-Quadruplex DNA at the Ensemble and Single-Molecule Level

Abstract

G-Quadruplex (GQ) DNA has garnered the attention of the scientific community due to its role in important biological processes like telomerase activity inhibition, regulation of gene expression and genomic stability. This has triggered studies on understanding the interactions of GQ with small molecules of therapeutic significance. We have studied the interactions of bioflavonoid kaempferol (3,4',5,7-tetrahydroxyflavone, KF), known for its anti-cancer properties, with a c-myc promoter-based GQ at the ensemble and single-molecule level. The binding constant (Kb) is estimated using steady state and time-resolved fluorescence measurements. Upon binding no structural perturbation of GQ is observable from the circular dichroism (CD) data. However, an increase in melting temperature of GQ upon binding suggests stabilization of the DNA structure by the ligand, an observation substantiated by the ensemble FRET experiments. Fluorescence correlation spectroscopy (FCS) measurements, which probe the interactions at the single-molecule level, reveal a progressive increase in the diffusion time of KF with increase in the amount of GQ confirming the binding event. The rate constants of the association (k+) and dissociation (k-) processes have been estimated from the FCS time profiles. The nature of the binding interactions and possible binding site of KF in GQ have been determined from the molecular docking studies. The above findings are expected to enhance our understanding of flavonoid-GQ DNA interactions for development of therapies involving quadruplex DNA.