TAV2b Peptide Derivatives Underwind and Stabilize Double-Stranded RNA upon Binding

Double-stranded RNA (dsRNA) has become an essential tool to understand biological processes with promising therapeutic implications. However, its usage is often limited due to poor cellular uptake and instability in biological settings. Peptidic dsRNA binders, inspired by natural RNA-binding proteins, have emerged as promising tools to address these limitations. However, it remains unclear how these peptides recognize RNA and impact its mechanical properties. Here we employed single-molecule magnetic tweezers to investigate TAV2b-derived peptidic dsRNA binders. We showed that these peptides underwind dsRNA upon binding and stabilize the resulting dsRNA conformation. Additionally, the wild-type peptide increases the dsRNA contour length while significantly lowering the persistence length. In contrast, a high-affinity homodimeric derivative condenses the dsRNA tether at forces below 1 pN. Furthermore, real-time experiments performed to understand the binding mechanism of TAV2b-derived peptides showed that the wild-type derivative is in dynamic association with dsRNA, whereas the homodimeric version forms a stable complex with dsRNA. Based on these findings, we propose a two-step equilibrium model where the RNA fluctuates between double-stranded and melted conformations, followed by peptide binding, which results in plectonemes. Our approach can inform the design of more potent and effective dsRNA binders for therapeutic and diagnostic applications.