Unraveling biofilm matrix composition, architecture, and function

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Bacteria are essential to life as we know it and colonize and thrive in astonishing environments, ranging from within the human host to the acidic microbial mats in Yellowstone and industrial oil pipelines. The propensity for bacteria to form multicellular communities termed biofilms far exceeds the tendency to persist in suspension. Bacteria secrete and surround themselves with molecular polymers as building materials to enmesh and protect resident bacteria in slime-like assemblies. The PI’s recent work revealed the assembly of extraordinary mechanically robust basket-like architectures surrounding E. coli cells in biofilms and led to the discovery of a new chemical structure never before observed in nature within these baskets. Cellulose is the most abundant biopolymer on Earth, present in the cell walls of plants and trees, leading to wood, paper, and cotton products. Other industries also employ methods to modify standard cellulose through synthetic routes for desirable properties and are used as food additives and in membrane and biotechnology applications. E. coli produces its own uniquely modified form of cellulose with phosphoethanolamine groups through newly defined molecular machinery. This project tackles crucial questions involving nature’s phosphoethanolamine (pEtN) cellulose: its modification patterning, its physical properties and potential for application in new industrial materials, and the molecular basis for the assembly of the full polysaccharide-protein nanocomposite structures. This project will fuel new research directions with implications for bacterial physiology; glycobiology; and industrial applications including the production of new materials and uses in renewable energy. The PI communicates scientific concepts and discoveries through scientific and educational research articles targeted to broad audiences. This project will train undergraduate and graduate students, particularly those from groups under-represented in STEM. The PI is designing coursework changes to introduce more quantitative concepts and hands-on course-based undergraduate research experiences (CUREs) into undergraduate chemistry courses.This project will integrate biochemical analysis with characterization of polymer strength and materials properties, electron microscopy, fluorescence microscopy, solid-state NMR spectroscopy and mass spectrometry to: (1) deliver a fundamental understanding of the molecular patterning and physical properties of the zwitterionic phosphoethanolamine cellulose; (2) determine the influence of the cellulose modification on the Velcro-like association of curli at the bacterial cell surface; (3) define atomic-level parameters corresponding to molecular interactions between curli and pEtN cellulose in native bacterial composites in situ and highly tuned complexes formed in vitro. In addition to revealing the fundamental chemical principles underlying biofilm matrix assemblies, the work will introduce new methods for the production of pEtN cellulose and isotopically labeled matrix materials, and provide unprecedented atomic- level analysis of insoluble matrix materials that pose a challenge to analysis by conventional methods. The project may serve to reveal a new paradigm for understanding the molecular basis of other amyloid- polysaccharide interactions, prevalent not only in microorganisms but also in eukaryotic cell systems and should inspire the search for alternately modified celluloses and polysaccharides. This project is supported by the Molecular Biophysics Cluster of the Molecular and Cellular Biosciences Division in the Directorate for Biological Sciences.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.