Time-Resolved Atomic Force Microscopy of Peptide and Protein Self-Assembly: Structure, Dynamics, and Mechanisms

Peptide and protein self-assembly is a common and important process in nature. Understanding the structural evolution and dynamic mechanisms of self-assembly is crucial for designing peptide materials and comprehending protein aggregation. Atomic force microscopy (AFM), with its high resolution and ability to perform in-situ imaging under near-physiological conditions, has become one of the core tools for studying the self-assembly behavior of proteins and peptides. This thesis focuses on the application of AFM in the study of peptide and protein self-assembly mechanisms. It comprehensively utilizes traditional AFM and high-speed AFM techniques, integrating them with other technologies to study systems ranging from simple to complex. The following three tasks were carried out: (1) Observing the self-assembly process of designed peptides transforming from a spiral conformation on surface to a fibril structure in solution, and comprehensively analyzing the driving factors of assembly using multiple methods. (2) Comparing the assembly behavior of α-helical and β-sheet peptides using AFM imaging, and exploring the regulatory effect of L- and D-conformation peptide mixing on assembly morphology. (3) Recording the dynamic assembly process of microtubules in situ using time-resolved AFM, and revealing the nucleation mechanism from bending to straightening by combining time resolved AFM with computational modeling. This study demonstrates the advantages of AFM in protein/peptide self-assembly research. The findings not only provide visual evidence for understanding the assembly mechanisms of specific peptide materials and proteins, but also offer a multi-scaled research framework for studying self-assembly mechanisms.