Fabrication of Tunable Mechanical Gradients by Mussels via Bottom-Up Self-Assembly of Collagenous Precursors

Functionally graded interfaces are prominent in biological tissues and are used to mitigate stress concentrations at junctions between mechanically dissimilar components. Biological mechanical gradients serve as important role models for bioinspired design in technically and biomedically relevant applications. However, this necessitates elucidating exactly how natural gradients mitigate mechanical mismatch and how such gradients are fabricated. Here, we applied a cross-disciplinary experimental approach to understand structure, function, and formation of mechanical gradients in byssal threads─collagen-based fibers used by marine mussels to anchor on hard surfaces. The proximal end of threads is approximately 50-fold less stiff and twice as extensible as the distal end. However, the hierarchical structure of the distal-proximal junction is still not fully elucidated, and it is unclear how it is formed. Using tensile testing coupled with video extensometry, confocal Raman spectroscopy, and transmission electron microscopy on native threads, we identified a continuous graded transition in mechanics, composition, and nanofibrillar morphology, which extends several hundreds of microns and which can vary significantly between individual threads. Furthermore, we performed in vitro fiber assembly experiments using purified secretory vesicles from the proximal and distal regions of the secretory glands (which contain different precursor proteins), revealing spontaneous self-assembly of distinctive distal- and proximal-like fiber morphologies. Aside from providing fundamental insights into the byssus structure, function, and fabrication, our findings reveal key design principles for bioinspired design of functionally graded polymeric materials.