Ezrin is a cytoplasmic protein that regulates epithelial cell function in multiple tissues and is also involved in invasion and metastasis of cancer cells. Ezrin can exist in closed (inactive) and open (active) conformational states, regulated by phosphorylation of T567 on its C-terminal domain. A T567D substitution mimics the open conformation, while a T567A mimics its closed form. Previous research suggests that the T567A ezrin cannot interact with its protein binding partners. However, we discovered that the T567A ezrin may directly interact with RNA, specifically those with guanine-rich regions that form G4 secondary structures (G4-RNA). We hypothesize that the binding of ezrin to G4-RNAs contributes to its metastatic phenotype. Therefore, investigating ezrin’s interactions with G4-RNAs is of paramount importance as this knowledge could lead to improved therapeutic strategies. Using molecular dynamics simulations, we examined molecular interactions of different RNAs with T567A ezrin. We used a thirty-five-nucleotide long RNA with five guanines in the middle, a stacked G4-RNA quadruplex having a bundle of four RNA copies, and one RNA unit from the bundled structure to create ezrin-RNA complexes. We analyzed simulation trajectories for these ezrin-RNA complexes to quantify binding affinities and to explore specific bonding. Our results showed favorable binding affinities with multiple hydrogen bonds responsible for the formation of these complexes. We confirmed the direct RNA binding using purified recombinant ezrin protein in surface plasmon resonance-based experiments and calculated a biding affinity (K D ) of ∼11.8 nM for this interaction. Our findings suggest that ezrin in its closed conformation may bind to RNA, which may have biologically relevant functions.
Tiwari et al. (Sun,) studied this question.