Grant: Project Grant - PA: Antimicrobial Resistance (Bridge Grant)

Many bacteria can form a dense matrix known as biofilm that helps them attach to surfaces and offers protection, aiding their survival. A key structural component of biofilm are amyloid fibrils, protein aggregates comprised of thousands of protein molecules assembled into nanofibrils that are 10-20 nm wide and 100-1000 nm long. Amyloid fibrils serve as a scaffold supporting the biofilm structure, and treatments targeting amyloid formation can be used to remove biofilm. The mechanism of amyloid formation is not well understood, but is a complex processes involving protein molecules changing shape, binding together, and assembling into specific fibrillar types-a process that is regulated in bacteria by various other proteins to inhibit or enhance amyloid formation. Here, we will use biophysical tools (optical tweezers and cryoEM) to resolve bacterial amyloid assembly from single molecules to the final mature fibrils, identifying key structural intermediates. Optical tweezers will be used to resolve the conformational changes of single protein molecules in order to map out the very first steps of protein self-assembly into fibrils, and how various other molecules regulate this process. The high-resolution structure of the mature fibrils will be solved using cryoEM. Understanding amyloid assembly at the molecular level will provide the structural basis for designing antibiofilm drugs.