Cholesterol Conjugated Elastin-like Recombinamers: Molecular Dynamics Simulations, Conformational Changes, and Bioactivity

Current models for elastin-like recombinamer (ELR) design struggle to predict the effects of nonprotein fused materials on polypeptide conformation and temperature-responsive properties. To address this shortage, we investigated the novel functionalization of ELRs with cholesterol (CTA). We employed GROMACS computational molecular dynamic simulations complemented with experimental evidence to validate the <i>in silico</i> predictions. The ELR^CTA was biosynthesized and characterized by using fluorescence assays, circular dichroism, dynamic light scattering, and differential scanning calorimetry. The <i>in silico</i> and <i>in vitro</i> data showed that CTA promotes the formation of intramolecular hydrogen bonds that favor β-sheet secondary structures. Compared with an unmodified ELR^VKV, CTA enhanced the hydrophobicity and stability of the system, allowing the formation of monodisperse nanoaggregates at physiologically relevant temperatures. Importantly, calorimetry assays revealed that ELR^CTA interacted and intercalated with the lipid bilayers of the DPPC liposomes. To demonstrate the implications of these changes for biomedical applications, ELR^CTA and DPPC-ELR^CTA hybrid nanoparticles were tested with cancer and immune cell lines. Interactions with the cell membranes demonstrated a synergistic effect of the composition and size of the modified recombinamer aggregates on the internalization. The results indicated the potential use of ELR-based nanoparticles for localized and systemic drug delivery. This work sets a new precedent to design elastin-inspired biomaterials with predictable self-assembly properties and develop novel drug delivery strategies.