Thermo-Responsive self-assembly of a dual glucagon-like peptide and glucagon receptor agonist

The discovery of functional amyloids has inspired tunable self-assembled materials composed of amyloid-like systems. Nanofibrils formed by the human peptide hormone Oxyntomodulin release active peptide after subcutaneous injection in rodents. Reversibility of Oxyntomodulin fibrillation is driven by destabilization of fibrillar forms at body temperature. This work provides the first evidence for amyloid-like depot formulation with optimum self-association forces below physiological temperatures. [Display omitted] •Oxyntomodulin reversibly self-assembles into amyloid-like nanostructures.•Reversible self-assembly could be used as a depot formulations.•Oxyntomodulin fibrillation kinetics and thermodynamics are sensitive to temperature.•Thermal sensitivity could trigger therapeutic monomer release from fibrils in vivo.•A mechanistic model for self-assembly and monomer release is suggested. The human peptide hormone Oxyntomodulin (Oxm) is known to induce satiety, increase energy expenditure, and control blood glucose in humans, making it a promising candidate for treatment of obesity and/or type 2 diabetes mellitus. However, a pharmaceutical exploitation has thus far been impeded by fast in vivo clearance and the molecule‘s sensitivity to half-life extending structural modifications. We recently showed that Oxm self-assembles into amyloid-like nanofibrils that continuously release active, soluble Oxm in a peptide-deprived environment. S.c. injected Oxm nanofibrils extended plasma exposure from a few hours to five days in rodents, compared to s.c. applied soluble Oxm. Here we show that Oxm fibril elongation kinetics and thermodynamics display a uniquely low temperature optimum compared to previously reported amyloid-like peptide and protein assemblies. Elongation rate is optimal at room temperature, with association rates 2–3 times higher at 25 °C than at ≥37 °C or ≤20 °C. We deduce from a combination of Cryo electron microscopy and spectroscopic methods that Oxm fibrils have a double-layered, triangular cross-section composed of arch-shaped monomers. We suggest a thermodynamic model that links the necessary molecular rearrangements during fibrillation and peptide release to the unique temperature effects in Oxm self-assembly and disassembly.