Symmetry-Guided Optical Control of Dynamic 1D Protein Self-Assembly and Precise In Vitro Initiation of Biomolecular...

Dynamic control over protein self-assembly into defined architectures remains an important objective in bionanotechnology. Here, we report a fully genetically encoded, photoreversible one-dimensional (1D) protein self-assembly system. By exploiting symmetry matching between D₂-symmetric StrepTactin and the photoswitchable tetrameric fluorescent protein Dronpa145N, we engineer a hierarchical 1D protein array whose assembly and disassembly can be reversibly toggled using specific wavelengths of light. Optical regulation of Dronpa145N oligomerization enables dynamic modulation of protein connectivity and effective valency without chemical cross-linkers or exogenous building blocks, providing a robust and modular platform for stimuli-responsive protein assembly. As an application of this controllable architecture, we demonstrate that light-induced array formation amplifies multivalency and can trigger liquid-liquid phase separation (LLPS) in vitro, whereas optical disassembly suppresses condensate formation. This system further enables precise temporal and spatial initiation of phase separation under well-defined conditions. Overall, this symmetry-guided, photoreversible assembly offers a versatile framework for engineering dynamic protein materials and serves as a phase-separating platform for dissecting the physicochemical principles underlying biomolecular condensate formation.