Chemical evolution of synthetic bacterial cells by reprograming protein translation with non-canonical amino acids

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The genetic code is one of the earliest manifestations of life. It is ubiquitous, unites all living organisms, and provides a shared biochemical basis for the entire biosphere. The long-term goal of my research is to understand the evolution and origin of the code, and to manipulate and expand the scope of protein biosynthesis. My experimental approach is directed towards construction of proteins with new and unique chemical functions, and manipulation of the chemistry of life as the result. This should provide an experimental ground for examination of evolutionary theories, and will enable construction of synthetic living cells. These goals are a shared endeavor of the worldwide community of scientists that extends beyond the borders of individual countries or scientific disciplines. The rationale of the research proposed under these two specific objectives is to answer fundamental scientific and technological questions and fill important gaps in the current state of knowledge. In the short term we have two major goals: I. Adaptive Laboratory Evolution (ALE) of Escherichia coli and Bacillus subtilis We will establish ALE as a method to propagate chemical changes in microbial populations. Specifically, ALE will be performed to introduce 3,4-dihydroxyphenylalanine (Dopa) globally into bacterial proteomes and single proteins in E. coli and B. subtilis species. Evolved strains will be used to elucidate biochemical mechanisms behind the adaptation. Final strains will be utilized for production of proteins endowed with unique features, such as adhesive capabilities or metal-binding. II. Tailor-made modular scaffolds designed by an expanded genetic code We will establish production of Dopa-containing proteins in microbial strains. An expanded genetic code approach will be used in order to bypass the posttranslational modification that is not readily available in prokaryotic microbial production strains. Dopa-containing proteins will be engineered based on modular scaffolds combining adhesive, elastic, and antimicrobial features. Their molecular properties such as metal coordinating, red-ox, acid-base, will be studied, and involvement of the Dopa residues in these features will be investigated. My newly established lab provides a state-of-the-art environment for trainees to learn the discipline of synthetic biology with opportunities for international and interdisciplinary collaboration with faculty in Chemistry, Microbiology and Bioengineering. Early-career researchers trained in multidisciplinary and multi-sectors research will acquire advanced skills for careers in industry, government, and academia. The proposed research will provide an experimental ground for the examination of evolutionary theories, and will enable construction of synthetic living cells for ultimate use in advanced biomaterials and technologies based on mussel adhesion proteins and lantibiotics (as a potential scaffold for the development of anti-infective agents).