Single-molecule magnetic tweezers reveals that TAV2b-derived peptides underwind and stabilize double-stranded RNA

Abstract

Double-stranded RNA (dsRNA) has evolved into a key tool in understanding and regulating biological processes, with promising implications in therapeutics. However, its efficacy is often limited due to instability in biological settings. Recently, the development of peptidic dsRNA binders derived from naturally occurring RNA-binding proteins has emerged as a favorable starting point to address this limitation. Nevertheless, it remains unclear how these high affinity dsRNA binders alter the structure and flexibility of dsRNA. To this end, we employed single-molecule magnetic tweezers experiments to investigate the effects of TAV2b-derived peptidic dsRNA binders on the mechanical properties of dsRNA. Torque spectroscopy assays demonstrated that these peptides underwind dsRNA, while also stabilizing the duplex. Additionally, force spectroscopy experiments demonstrate that a wild type TAV2b peptide derivative extends the contour length and lowers the bending rigidity of dsRNA, while a homodimeric version triggers the formation of higher order complexes at forces below 1 pN. Our study presents a quantitative approach to investigate how these peptides alter the structure of dsRNA, and whether peptide structural design alters the affinity to dsRNA and its stability. This approach could inform the design of more potent and effective dsRNA binders in the efforts to advance RNA therapeutics.