A Multimodal Lens on Protein Assembly : From Silk to Seeds

Protein-based materials rely on controlled assembly across multiple lengthscales. Key organisational features often emerge before macroscopic fibres orgels form. Yet the pathways linking molecular organisation to material formationremain poorly resolved. This is partly because structural evolution across lengthscales is rarely measured concurrently within the same sample.This thesis examines protein assembly as a multiscale, pathway-dependentprocess. Silk proteins serve as the primary model system, as they form highlyorganised fibres under mild aqueous conditions. Yellow pea proteins provide acomplementary multicomponent case in which processing history and co-existingcomponents influence assembly.The work focuses on the solution and pre-gelation states. In reconstitutedsilk fibroin from Bombyx mori, time-resolved multimodal measurements are used.Neutron scattering is combined with ultraviolet and fluorescence spectroscopyto follow structural evolution from molecular to mesoscopic scales within thesame samples. These measurements reveal structured intermediate states priorto extensive β-sheet formation. The accessibility and persistence of theseintermediates depend on the assembly environment, showing that processingconditions select assembly pathways rather than simply triggering final states.Native and recombinant cylindriform spidroins are examined as a simplerspider silk system. Small-angle X-ray scattering and ensemble modelling revealelongated, modular solution architectures, linking solution-state organisation toits multi-domain architecture. Recombinant constructs further demonstrate thesensitivity of assembly to junction stability and solution context.In pea protein–starch mixtures, contrast-variation neutron scattering shows thatproteins and residual starch are largely independent in solution. Thermal treatmentinduces co-assembly in a processing-history-dependent manner.Together, these studies show that protein-based material formation is shapedupstream of macroscopic assembly. Resolving organisation across length scaleshighlights the roles of pre-organisation, pathway-dependent intermediates,and solvent participation. The multimodal and scattering-based approachesestablished here provide a general strategy for investigating hierarchical proteinassembly in both silk-like and food-relevant systems.