Confinement-Induced Self-Assembly of Protein Nanofibrils Probed by Microfocus X-ray Scattering

Area of Science:

• Biophysics
• Materials Science
• Protein Self-Assembly

Background:

• Whey protein nanofibrils (PNFs) are self-assembling protein structures with potential applications in biomaterials.
• Understanding the factors influencing their assembly into larger structures is crucial for controlling material properties.

Purpose of the Study:

• To investigate the confinement-induced assembly of whey protein nanofibrils (PNFs) into microscale fibers.
• To explore the role of temperature, nanofibril morphology, and flexibility on the assembly process and resulting structures.

Main Methods:

• Microfocused synchrotron X-ray scattering was used to study the in situ assembly of PNFs within a droplet.
• Solvent evaporation was employed to induce alignment and fiber formation.
• The influence of nanofibril persistence length and flexibility was examined.

Main Results:

• An optimal temperature was identified that maximizes the order parameter of the protein fibers.
• Nanofibril morphology (stiff/straight vs. flexible/curved) significantly impacts assembly behavior and final structure.
• Stiff PNFs showed interface alignment and central entanglement, while flexible PNFs exhibited more uniform alignment and network formation.

Conclusions:

• The degree of order in protein fibers is a balance between forced alignment and rotational diffusion, influenced by temperature.
• Nanofibril flexibility and nanoscale morphology dictate assembly pathways, leading to distinct microscale fiber architectures.
• This study provides insights into controlling protein self-assembly for tailored biomaterial design.