The BIO-SENS program aims to develop engineering principles for on-demand protein biosensors. As part of BIO-SENS, we worked toward developing components of analyte-responsive protein polymers, consisting of interleukin-6 (IL-6) binding mini-proteins for analyte recognition and stimuli-responsive elastin-like polymers (ELPs) for signal transduction. We expressed and purified recombinant IL-6 binders from a bacterial expression system using affinity chromatography, optimizing protocols to improve quality and reproducibility. To characterize ELPs, we integrated UV-Vis spectrophotometry and dynamic light scattering temperature-ramping experiments, refining experimental parameters and buffer conditions to capture phase-transition behavior. We compared the kinetics of phase transition in response to salt of two ELPs, termed I40 and V40, which differed in composition and polarity. Our results revealed distinct kinetics between the two ELP variants, with the more hydrophilic V40 undergoing salt-induced phase transition more rapidly and with a different assembly profile than the more hydrophobic I40, demonstrating the sensitivity of polymer behavior and environmental responsiveness to sequence-specific properties. These findings provide insight into the fundamental behavior of ELP-based systems and establish improved methods for reproducible protein purification and analysis. Further studies will extend these methods by testing ELP behaviors with a broader range of stimuli and contexts, including on sensor surfaces.
Falebita et al. (Sun,) studied this question.