Local changes in individual spider silk fibers from tubuliform and major-ampullate glands under varying humidity and tensile strain

Spider silk’s exceptional mechanics and environmental adaptability stem from a hierarchical structure of crystalline β-sheet nanodomains embedded in an amorphous protein matrix. While major ampullate (MA) silk’s structural responses to strain and environment are well studied, tubuliform (TU) silk—critical for egg sac construction and promising for biomedical uses—remains poorly understood. We investigate TU and MA silk under varying relative humidity (RH) and mechanical strain using spatially resolved, high‑resolution nanobeam X‑ray diffraction. Increasing RH markedly reduces Young’s modulus, yield strain, and yield stress in both silks, with TU showing greater humidity sensitivity. Ultrastructural analysis reveals key contrasts: TU exhibits pronounced changes in nanocrystal dimensions, molecular alignment, and lattice spacing with strain and humidity, whereas MA maintains more stable crystalline organization, reflecting optimization for tensile strength. By mapping structural parameters across single-fiber diameters (edge, transition, bulk), we provide spatially resolved insights into TU’s ultrastructure and hierarchical mechanisms. Notably, structural parameters are predominantly homogeneous across the fiber diameter for both silks at the studied length scales. These distinct structural responses offer design templates for biomimetic materials. Future work should examine additional environmental variables (e.g., temperature, solvents) and finer length scales to further unlock spider silk’s potential as a model biomaterial.