BPS2026 – Thermodynamic influence of cationic cluster size on protein-assisted nucleic acid folding

Nucleic acid-binding proteins (NABPs) help NAs adopt complex secondary and tertiary structures by lowering energetic barriers to folding through transient or specific interactions. This work combines single-molecule FRET experiments with coarse-grained molecular dynamics simulations to assess the fundamental principles that govern protein-assisted NA folding. Specifically, we explore how clustering of cationic residues in NABPs influences a four-state conformational equilibrium model of a NA secondary structure by systematically “re-building” NABPs from their smallest cationic constituents (e.g., ammonium→ Lys→ polyK 10 → NABP). The results of these efforts show that the size of cationic clusters in NABPs contributes to multiple aspects of protein-assisted NA folding, such as binding affinity and structural compaction but does not dictate folding favorability. Furthermore, the results of our coarse-grained molecular dynamics reveal novel structural and mechanistic insights pertaining to the number and location of bound NABPs.