Free energies of bimolecular associations and the mechanisms of nanoscale molecular self-assemblies

Understanding how ultrashort peptides initiate self-assembly requires clarifying the earliest bimolecular association events. Here, we investigate three hydrophobic tripeptides—FFF, LLL, and III—to determine how their molecular geometry governs dimer formation and subsequent assembly pathways. Two-dimensional free-energy landscapes obtained from umbrella sampling reveal a universal preference for antiparallel alignment but sharply different propensities for forming middle-to-middle hydrogen bonds. III readily forms these stabilizing interactions, LLL forms them partially, while FFF is strongly disfavored. Long unbiased simulations confirm that stable dimers contribute significantly to the growth of the final aggregate in III, moderately in LLL, and minimally in FFF. Event-resolved analyses further show that dimer fusion is a dominant growth mode for III but negligible for FFF. Together, these results establish middle-to-middle hydrogen bonding as the key determinant distinguishing the self-assembly mechanisms of ultrashort peptides, providing a predictive framework for designing peptide-based supramolecular materials.