Compression‐Driven Self‐Assembly of Fusion Peptides Into Fibrous Networks Revealed by In Situ Grazing‐Incidence X‐Ray Diffraction

Fusion peptides (FPs) have emerged as versatile building blocks for nanoscale architectures with potential applications in nanolithography, surface templating, and bioinspired materials. However, most studies of FP self-assembly at fluid interfaces rely on ex situ characterization after transfer to solid substrates, making it difficult to determine whether the observed structures form at the interface or during transfer and drying. Here, we employ grazing-incidence X-ray diffraction (GIXD), combined with X-ray reflectivity (XRR), to directly probe in situ the structural evolution of FP monolayers at the air-water interface during uniaxial compression. The results reveal that compression drives the development of lateral molecular order and fibrillar assemblies at the fluid interface. For FP1, increasing surface pressure promotes β-sheet ordering and fibril coherence, with maximum in-plane correlation at an intermediate pressure, whereas further compression partially disrupts this organization. In contrast, FP4 exhibits only short-range order with limited response to compression. These findings identify surface pressure as a physical parameter for controlling peptide organization at fluid interfaces and provide a basis for compression-directed fabrication of tunable peptide nanostructures.