Engineered β-Crystal Domains Enable Strong Humidity-Responsive Actuation in Recombinant Spider Silk

Designing humidity-responsive protein fibers that combine high recovery stress with structural integrity is essential for advancing soft actuators under physiological conditions. However, conventional polymer-based actuators are limited by low mechanical strength and poor humidity tolerance. Spider silk provides a natural model for water-responsive actuation, yet replicating its performance in recombinant systems remains challenging due to hydration-induced β-sheet disruption and insufficient crystalline stabilization. Here, recombinant spidroin fibers are engineered by introducing terminal cysteine crosslinking, enabling site-specific disulfide bonds to form during shear-assisted wet spinning. This covalent edge reinforcement preserves β-sheet alignment even at 90% relative humidity, as confirmed by molecular dynamics simulations and spectroscopic analyses. The optimized C4S fibers exhibit reversible and controllable humidity-driven actuation, delivering rapid contraction with a recovery stress of 45 MPa and a work density of 122 kJ m⁻³, exceeding typical synthetic actuators and surpassing human skeletal muscle by over threefold. This sequence-encoded crystalline locking strategy provides a generalizable molecular design for creating moisture-resilient, high-performance protein actuators, with potential applications in soft robotics, adaptive textiles, and biomedical devices. © 2026 Wiley-VCH GmbH.