(164r) The Influence of Electrostatic Distribution on Small Molecule Binding of Coiled-Coil Protein Microfibers

The ability to engineer the solvent-exposed surface of self-assembling coiled-coils allows one to achieve higher order hierarchical assembly such as nano or microfibers. Currently these materials are being developed for a range of biomedical applications, including drug delivery systems, however ways to mechanistically optimize the coiled-coil structure for drug binding has yet to be explored. Our laboratory has previously leveraged the functional properties of the naturally occurring cartilage oligomeric matrix protein coiled coil (COMPcc), not only for its favorable motif but also for the presence of a hydrophobic pore to allow for small molecule binding. Here, we present a small library of protein microfibers derived from the parent sequence of COMPcc with various electrostatic potentials, with the aim to investigate the influence of higher order assembly on candidate small molecule, curcumin, drug delivery. The impact of drug binding on coiled-coil mutants was characterized by fluorescence spectroscopy and confocal microscopy. Structural characterization was also performed via circular dichroism and attenuated response Fourier transform infrared spectroscopy. Our findings demonstrate that redistribution of the surface charge impacts not only its supramolecular assembly but also its thermal stability after drug binding. We elucidate this relationship with the support of Rosetta docking simulations, revealing the impact of surface charge on side-chain packing in the pore. For all variants, protein melting temperature saw a modest to extreme improvement upon small molecule encapsulation. By continuing to investigate these structure-property-function relationships, we can construct electrostatically optimized protein fibers capable of efficiently encapsulating and stabilizing small molecules at a higher rate than the current state-of-the-art coiled coil drug delivery system.