Resolve Protein Two-Dimensional Dynamics at Solid-Liquid Interfaces with High-Speed Atomic Force Microscopy

Proteins, the fundamental building blocks of nature, possess an inherent ability to self-assemble into intricate hierarchical structures with diverse functional properties. Recently, scientists have started leveraging this self-assembly capability to create innovative biomimetic materials for applications in health, energy, and environmental sectors. Despite these advances, the controlled fabrication of two-dimensional (2D) protein architectures at solid-liquid interfaces remains a significant challenge. The dynamics and ultimate organization of proteins at these interfaces are governed by the complex energy landscape arising from the interplay between protein-protein and protein-surface interactions, which is not yet fully understood. To unravel how this energy landscape responds to varying solution conditions and external stimuli at solid-liquid interfaces, in-situ characterization techniques with high temporal and spatial resolution and comprehensive statistical analysis are essential.
Recently, we developed strategies to guide protein assemblies at inorganic interfaces through carefully programmed protein-protein and protein-substrate interactions. In this presentation, I will discuss the assembly behavior of DHR10-MicaN, protein nanorods that form distinct 2D matrices in response to varying KCl concentrations and the properties of the mica interface [1]. We employed high-speed AFM to capture the in-plane dynamics of the protein nanorods on mica as a function of KCl concentration. The resulting data were then analyzed using a deep learning approach specifically adapted for AFM data involving rod-shaped objects [2, 3, 4]. We quantitatively described the in-plane dynamics of the protein nanorods, which are influenced by DHR10-MicaN-mica binding affinity, entropic effects, interfacial hydration layers, and substrate symmetries [3, 5, 6]. These findings reveal the extensive range of self-assembled architectures formed by these protein nanorods; exceeding expectations based on their basic design. More broadly, this study establishes a methodology for understanding how cations and engineered protein interfaces govern the in-plane dynamics of macromolecules at solid-liquid interfaces.

References:
1. Pyles, H., et al., Nature 571 (2019), 7764
2. Kalinin, S.V., et al., ACS NANO 15 (2021), 6471
3. Zhang, S., et al., Proceedings of the National Academy of Sciences 119 (2022), e2020242119
4. Ziatdinov, M., et al., Nano Letters 21 (2021), 158
5. Alberstein, R.G., et al., Journal of Physical Chemistry Letters, 14 (2023), 80
6. Prelesnik, J.L., et al., Proceedings of the National Academy of Sciences, 118(2021), e2025121118.