Topological features of the intermolecular contacts in gluten-forming proteins: Exploring a novel methodological...

This work introduces a novel methodological approach to study both the geometry of complex protein networks and the nature of the interacting proteins. This approach is based on the high reactivity of Au⁺ ions on the surface of gold nanoparticles (AuNPs) towards thiols, that allows fast formation of a covalent bond between accessible protein thiols and AuNPs. In the case of the durum wheat semolina used in the exploratory studies reported here, the nature of proteins covalently bound to AuNPs is expected to be affected by both the compactness of the protein network and by the AuNPs size. Simple centrifugation procedures allowed recovery of the protein-loaded AuNPs that remained soluble, and the protein(s) covalently bound on the surface of soluble AuNP were identified by MS analysis of their proteolytic fragments. Gluten-forming proteins were found to be bound to soluble AuNPs only when detergents or chaotropes were added to the semolina/AuNPs suspension at room temperature. AuNPs-bound proteins also included gluten-forming proteins with no reported free thiols, suggesting that they are piggybacked on other thiol-containing gluten-forming proteins via disulfide bonds already present in the otherwise untreated semolina. The potential of this approach is discussed in terms of the possibility of developing a methodology suitable for further clarification of the geometrical features of protein networks, of the nature of the involved proteins, and of the type of interaction they establish, as well as any modifications of these features upon processing.