The GCE4All Center: Unleashing the Potential of Genetic Code Expansion for Biomedical Research

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The proposed GCE4All Biomedical Technology Development and Dissemination Center at Oregon State University (OSU) will optimize, develop, and broadly disseminate Genetic Code Expansion (GCE) technology – the engineering of cellular translation to express proteins containing non-canonical amino acids (ncAAs). GCE provides unprecedented ways to probe and manipulate macromolecular structure and function, analyze protein malfunctions in disease, engineer bioanalytical tools, and create new precision biotherapeutics. GCE's feasibility is well-established, but it remains difficult for researchers to access and implement, and thus remains little-used – an ideal target for BTDD support. During its envisioned lifespan of ≤15 years, the Center's mission will be to optimize and extend existing GCE technologies to enable facile use by non-specialists, and to broadly disseminate them via widespread education, effective training, and by providing sustainable access to optimized technologies via established repositories – enabling powerful GCE approaches to become standard, widely-used tools of biomedical researchers. Advantageous for creating the proposed Center is our experience and leadership in the groundbreaking predecessor OSU Unnatural Protein (UP) Facility (2012-21), which at a much smaller level developed and disseminated GCE methods and trained researchers. GCE4All Center leaders are thus well- equipped to accomplish the Center mission via its 2 Technology Development Projects (TDPs), 10 initial Driving Biomedical Projects (DBPs), and Community Engagement (CE). The synergistic TDPs will optimize and extend GCE methods for 1) bioorthogonal ligation applications using GCE-produced proteins, including low- background labeling and tracking in mammalian cells, and 2) producing ncAA-proteins that contain biochemical probes and/or native or analog post-translational modifications (PTMs) – ubiquitous but little-understood regulators of protein functions. To ensure broad relevance, targeted technology advances will overcome barriers faced by geographically-diverse, NIH-funded DBPs that will serve as stringent testbeds for the work. To achieve its “GCE for All”goal, Center optimizations will bridge 4 common technological barriers that deter researchers from adopting GCE. These 4 GCE Bridges include: effective tools for 1) incorporating ncAAs of choice; for efficiently producing impurity-free GCE proteins in 2) E. coli and 3) mammalian cells; and 4) creation of stable mammalian cell lines and reliable protocols for reproducible studies in cells. The CE core will provide diverse training activities including hands-on workshops already proven effective by our UP Facility. Via the Center website, CE will disseminate GCE methods, online training, and host a Wiki GCE-knowledgebase and a GCE4All community networking bulletin board enabling peer-to-peer support in the GCE user community. The CE core will also organize biennial International GCE Conferences and ensure all optimized reagents are publicly available from repositories. The GCE4All Center will achieve these ambitious goals in Years 1-5, toward its ultimate goal of transforming GCE from a boutique method to a standard part of the molecular biologist's toolkit.