Temperature‐triggered phase separation of recombinant proteins has offered substantial opportunities in the design of nanoparticles for a variety of applications. Herein, the temperature‐triggered phase separation behavior of a recombinant hydrophilic resilin‐like polypeptide (RLP) is described. The transition temperature and sizes of RLP‐based nanoparticles can be modulated based on variations in polypeptide concentration, salt identity, ionic strength, pH, and denaturing agents, as indicated via UV–Vis spectroscopy and dynamic light scattering (DLS). The irreversible particle formation is coupled with secondary conformational changes from a random coil conformation to a more ordered β‐sheet structure. These RLP‐based nanoparticles could find potential use as mechanically‐responsive components in drug delivery, nanospring, nanotransducer, and biosensor applications.
