Cryo-ET Analysis Unveils the Minimum Structural Unit and Hierarchical Ultrastructure of Silk Fibers in Silkworm and Spider

Silkworm silk and spider silk are renowned for their exceptional mechanical properties, with the toughness of spider dragline silk exceeding that of silkworm silk and even outperforming high-performance synthetic fibers such as Kevlar and nylon. The mechanical properties of silk fibers are determined by their ultrastructure, including the size, morphology, and arrangement of their fundamental structural units. In this study, we employed the state-of-the-art cryo-electron tomography to investigate the in situ ultrastructure of silkworm silk, spider silk, and artificial silk. Our findings reveal that the fundamental structural unit of silkworm silk is a nanofibril approximately 3.6 nm in diameter, connected by numerous bridges, representing the smallest structural unit currently observable. These nanofibrils are aligned parallel to the fiber axis and organized into a herringbone pattern oriented in a specific direction. Multiple layers of this herringbone pattern stack together to form the final micron-scale fiber. In spider silk, a dense structure featuring nanofibrils is clearly visible, with their long axes tightly aligned parallel to the flow direction, leaving no discernible gaps. By contrast, silkworm silk exhibits larger, regionally varying gaps between nanofibrils, while artificial silk lacks the highly ordered arrangement characteristic of natural silks. Our study provides new insights into silk formation and offers valuable principles for designing ultrastrong silk.