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

Abstract 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.