UC Santa Barbara

The desirable physicochemical properties and functions of membrane proteins (such as catalysis, biosensing, and ion transport) have generated interest for guest protein moieties in abiotic host systems for technological applications. Specifically, I have used a self-assembled nanostructured silica material host system to exploit the functionality of the light-activated H+ ion-pump proteorhodopsin outside of its native cell membrane. Incorporated into a device, this protein is promising for applications as a sustainable bio-based alternative for solar-to-electrochemical energy conversion. Traditionally, the inclusion of membrane proteins into non-native inorganic materials has been challenging, in part because the surfactants, polymers, organic solvents, and synthesis conditions used to create well-ordered nanostructures often cause the protein to denature, whereas the biocompatible surfactants that are capable of stabilizing membrane proteins often do not produce the well-ordered nanostructures required of devices. Judicious selection and combination of structure-directing and protein-stabilizing short-chain nonionic surfactants and charged phospholipids has allowed us to stabilize membrane proteins in the cylindrical hydrophobic regions of surfactants within silica-surfactant materials with high extents of mesostructural order. Furthermore, by optimizing the composition of the host film, I have achieved a record high 44 wt% concentration of functionally active guest membrane proteorhodopsin, which improve the prospects for macroscopic proton transport in devices.

Techniques were drawn from multiple fields of study to characterize and understand how these non-native host materials self-assemble, and how they impact the photocycle kinetics of proteorhodopsin guest molecules. These techniques include small-angle X-ray scattering (SAXS) to determine the mesoscale structure of the silica film, solid-state nuclear magnetic resonance (NMR) spectroscopy to analyze the atomic-scale composition and structure at the surfactant-silica interface, nanofabrication processes to alter substrates and direct self-assembly at the film-substrate interface, cryo-electron microscopy (cryo-EM) to determine the structure and intermolecular interactions of proteorhodopsin in precursor micellar assemblies, and time-resolved UV-visible light spectroscopy to quantify differences in the photocycle of proteorhodopsin due to differences in local host environments. Critically, these nanostructured protein-surfactant-silica films are well-ordered and display promising protein loading, mechanical and thermal robustness, and protein stabilizing effects necessary to exploit the highly desirable functions of membrane proteins. A novel and interdisciplinary approach to characterization, understanding, and subsequent optimization of proteorhodopsin and nanostructured silica films has enabled the preparation of robust abiotic inorganic-organic host materials with record high concentrations of functionally active guest membrane proteins, a significant step towards the integration of membrane proteins into devices.