CAREER: Post-translationally Lipidated Biopolymers As Multiphasic All-Aqueous Emulsions

Syracuse University, Syracuse NY

Investigators

Abstract

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2). PART 1: NON-TECHNICAL SUMMARY Emulsions are suspensions of two immiscible liquids, which are ubiquitously utilized across many industries for diverse applications. The research goal of this CAREER project is to synthesize biomaterial-based water-in-water emulsions using a process inspired by natural cell biology. Cells create such emulsions by leveraging phase separation of their intracellular proteins and have developed capabilities to tailor the properties of these emulsions by modifying the phase separating proteins using a diverse palette of chemical reactions. Our understanding of how these modifications stabilize or tailor the water-water interfaces remains incomplete, which increases our reliance on traditional oil-based emulsions. To address this challenge and advance the field of biomaterial design, this research uses a combination of genetic and chemical manipulation to synthesize and study phase-separating proteins with customizable modification patterns. The size and stability of resulting emulsions will be characterized and will be correlated to the modification patterns. The knowledge gained from these studies will be used to create water-in-water emulsions that can be precisely controlled to have user-defined characteristics. Due to their biocompatibility, such new water-in-water emulsions have great potential to replace traditional oil-based emulsions in food processing, cosmetics, biosensing, and delivery of pharmaceuticals. This project will provide the materials science community with innovative and easy-to-use platforms for scalable synthesis of biomaterials and help accelerate the ability to manufacture the next generation of structurally complex, functionally tunable biomaterials for multiple industrial and healthcare applications. This will strengthen the U.S.'s leadership in the global bioeconomy. Educational activities supported through this project will train the next generation of scientists and engineers in bio-enabled technologies. A major educational activity is the student creation of biodegradable and eco-friendly materials for fabric and textile applications. Applications of this work can address the highly polluting and environmentally damaging current practices in textile manufacturing. This educational activity will be used in outreach to attract and engage high school students to this merger of science and design. In addition, this project will inform the redesign of an undergraduate and graduate laboratory course that will involve students in real-world discovery and will train students in the regulatory framework used in the biotechnology industry. PART 2: TECHNICAL SUMMARY This CAREER project will exploit the bio-enabled process of post-translational modification (PTM) to advance the goal of rationally designing hybrid biomaterials with functionality that exceeds the capabilities of natural biopolymers. The study will decrypt the material-design principles of phase-separating lipidated proteins (PLPs) to create water-in-water emulsions with user-defined formation, fluid properties, and internal structure. The study will investigate the overarching hypothesis that the physicochemical interplay between the protein, lipid, and lipidation site domains regulates the strength and half-life of adhesive and cohesive interactions that control the formation, viscoelasticity, and hierarchical organization of PLP condensates. Three research objectives will elucidate the molecular and thermodynamically grounded descriptions of how lipidation PTM alters the interfacial and rheological characteristics of model protein condensates: (1) correlate the thermodynamic stability of PLP emulsions to the physicochemistry of lipid and phase-separating proteins, (2) ascertain the effect of lipidation site sequence on the dynamics of cohesive interactions and colloidal properties of biphasic PLP emulsions, and (3) determine the molecular factors that regulate the strength of adhesive microscopic interactions and macroscopic miscibility of multiphasic PLP emulsions. Methods to achieve these objectives include bio-/semi-synthesis of PLPs to create libraries with systematically varied lipid, protein, and lipidation sites; determination of phase boundaries using light scattering techniques; and characterization of dynamic material properties of PLP droplets using microscopy and phase-separated states using rheology. Multidimensional nuclear magnetic resonance techniques will be used to investigate the defining structural and dynamic elements of PLPs to enable the rational design of the next generation of condensates with tailorable structural and material properties. The education plan integrates these research efforts into a multitiered approach to (1) increase the public's awareness of bio-enabled materials and technologies as solutions to societally relevant problems, (2) recruit students from nontraditional backgrounds to biomaterials research, and (3) enhance biomaterials education at the undergraduate and graduate levels. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.