Enhanced yields and purity: a robust optimization framework for the production of multifunctional elastin-like proteins.

Elastin-Like Proteins (ELP) are highly versatile biomaterials with significant potential in tissue-engineering, due to their design flexibility, mechanical properties and tunable stimuli responsiveness. However, expression conditions reported in literature do not always translate to new ELP fusion proteins. This study establishes a methodology to optimize expression conditions for modular ELP constructs with intermittent peptide domains. Response Surface Methodology is employed to systematically optimize protein expression of three distinct ELP constructs (containing either RGD domains, heparin-binding domains, or a combination thereof) currently being explored in the domain of injectable biomaterials. By combining small-scale expression with western blot densitometry, an accessible workflow is established to systematically optimize expression yields while avoiding extensive sample preparation and limiting the number of experiments. Analysis of the lower critical solution temperature of the ELPs suggests a correlation between the molecular architecture and the observed transition temperature. Furthermore, in vitro biocompatibility analyses confirm high cell viability, validating these modular ELPs as promising candidates for injectable hydrogels.