pH-triggered clustering regulates β-sheet activation in silk assembly

Area of Science:

• Biomaterials Science
• Protein Chemistry
• Materials Science

Background:

• Silk fibers possess unique properties due to their hierarchical protein organization.
• The molecular mechanisms governing silk fibroin's structural transformation are not fully understood.

Purpose of the Study:

• To elucidate the stepwise assembly pathway of regenerated silk fibroin gelation.
• To investigate the role of gradual acidification versus rapid aggregation in silk structure formation.
• To define the sequence and timing of events in silk's hierarchical architecture construction.

Main Methods:

• Time-resolved small-angle neutron scattering (TR-SAXS).
• Turbidity and fluorescence emission measurements (NUrF).
• Comparative analysis of biomimetic gradual acidification and methanol-induced gelation.

Main Results:

• Identified a stepwise assembly pathway: nanoscale clustering, domain growth, and mesoscale network formation during gradual acidification.
• Observed unique intermediates and a regulated onset of β-contacts and β-sheets assembly.
• Methanol-induced gelation bypassed these intermediates, leading to rapid aggregation.

Conclusions:

• Silk fibril formation requires prior compaction and network connectivity, achieved through regulated assembly.
• Pathway selection critically governs the material outcomes in protein self-assembly.
• The NUrF technique offers a broadly applicable strategy for studying hierarchical assembly in protein materials.