Protein function evolution through the lens of conformational dynamics: A single-molecule perspective.

Abstract

Understanding how proteins evolve requires moving beyond the classical sequence-structure paradigm toward a framework that incorporates conformational dynamics as a substrate of innovation. Within this framework, function emerges not only from single structures but from an ensemble encoded in the sequence. In this context, single-molecule Förster resonance energy transfer, which resolves heterogeneous and transient conformations inaccessible to ensemble methods, has shown that dynamics can tune activity, specificity, and regulation. Importantly, disordered proteins - which maintain biochemical properties rather than precise sequences - represent the limiting case of this principle, where dynamics become the primary substrate for evolution. Recent advances have extended these principles to protein design, demonstrating that dynamics are not only evolvable but also programmable. Together, these insights position conformational dynamics at the center of understanding and engineering the evolutionary logic of protein function, opening the door to study how proteins are tuned to operate under the nonequilibrium conditions of living cells.