RNA homodimerization strand displacement pathways to extended duplexes: Atomistic details for a mechanistic paradigm to identify unique antiviral targets for current and emerging viral pandemics - Project Summary Kissing complexes (KC) and extended duplexes (ED) represent dynamic RNA structures whose interconversion is an example of strand displacement reactions that has been implicated in playing vital roles in several viruses, influencing a broad spectrum of biochemical processes including genome packaging, viral recombination, and host-pathogen interactions. However, the structural, dynamic, and energetic details on how these interconversions occur are virtually unknown yet are vital in our novel approach of identifying and characterizing previously unconsidered meta-stable pathway states to inspire future antiviral and drug resistance development. Overcoming significant challenges that hamper meaningful modelling of RNA and dimers, and associated interconversion pathways necessitate judicious choice of computational techniques and integration with experimental data across physiologically relevant timescales. Our overarching hypothesis is that by leveraging the limited kinetic and thermodynamic data reported on RNA transition pathway systems, including the human immunodeficiency virus 1 (HIV-1) dimer initiation site (DIS) and model complexes based on the bacterial E. coli DsrA-rpoS RNA-mRNA regulatory complex, a general paradigm will be developed for other known RNA dimerization systems, such as SARS-CoV, SARS-CoV-2, HCV, and future emerging RNA viruses requiring novel antiviral therapies. To address our hypothesis, we first employ a unique approach to establish the unbiased structure and dynamics of RNA structures representing both pathway endpoints of each strand displacement reaction, such as KC to ED (Aim I). Confidence in structural predictions for systems without experimental structures is engendered by comparing computations against HIV-1 DIS crystallographic structures. We next apply a minimum energy pathway technique with and without protein chaperones to identify meta-stable states (Aim II). Reliability of the method is evaluated against the kinetic and thermodynamic data reported for the KC to ED interconversion for the E. coli DsrA-rpoS RNA-mRNA regulatory complex. Finally, as a proof of concept, we will screen RNA-binding molecules and antisense oligonucleotides against identified meta-stable intermediates along the pathways (Aim III). Aligned with NIH/R15 goals, we will train undergraduates to understand the capabilities, limitations, and errors of experimental and computational chemistry to craft ultimately a seamless research approach. Our training objective is to deliver a quality research experience that motivates undergraduates to achieve their highest potential and best prepare them for scientific research and discovery. The intent is to attract and retain the nation’s diverse student talent pool, having the consequence of enriching and diversifying the U.S. workforce by adding experts in the field of biomedical chemistry. The expected scientific outcomes are to unlock novel physical insights into strand displacement reactions and provide a foundation for targeted drug design and therapeutic interventions for drug resistance of HIV-1, SARS-CoV2, and HCV viruses of current
