Many supramolecular materials in biological systems are driven to a nonequilibrium state by the irreversible consumption of high-energy molecules such as ATP or GTP. As a result, they exhibit unique dynamic properties such as a tunable lifetime, adaptivity or the ability to self-heal. In contrast, synthetic counterparts that exist in or close to equilibrium are controlled by thermodynamic parameters and therefore lack these dynamic properties. To mimic biological materials more closely, synthetic self-assembling systems have been developed that are driven out of equilibrium by chemical reactions. This protocol describes the synthesis and characterization of such an assembly, which is driven by carbodiimide fuels. Depending on the amount of chemical fuel added to the material, its lifetime can be tuned. In the first step, the protocol details the synthesis and purification of the peptide-based precursors for the fuel-driven assemblies by solid-phase peptide synthesis. Then, we explain how to analyze the kinetic response of the precursors to a carbodiimide-based chemical fuel by HPLC and kinetic models. Finally, we detail how to study the emerging assembly’s macro- and microscopic properties by time-lapse photography, UV-visible spectroscopy, shear rheology, confocal laser scanning microscopy and electron microscopy. The procedure is described using the example of a colloid-forming precursor Fmoc-E-OH and a fiber-forming precursor Fmoc-AAD-OH to emphasize the differences in characterization depending on the type of assembly. The characterization of a precursor’s transient assembly can be done within 5 d. The synthesis and purification of a peptide precursor requires 2 d of work. © 2021, The Author(s), under exclusive licence to Springer Nature Limited.     «

Many supramolecular materials in biological systems are driven to a nonequilibrium state by the irreversible consumption of high-energy molecules such as ATP or GTP. As a result, they exhibit unique dynamic properties such as a tunable lifetime, adaptivity or the ability to self-heal. In contrast, synthetic counterparts that exist in or close to equilibrium are controlled by thermodynamic parameters and therefore lack these dynamic properties. To mimic biological materials more closely, syntheti...     »