Snapshots of usher-mediated protein secretion and ordered pilus assembly

Type 1 pilus biogenesis was used as a paradigm to investigate ordered macro molecular assembly at the outer cell membrane. The ability of Gram-negative bacteria to secrete proteins across their outer membrane and to assemble a dhesive macromolecular structures on their surface is a defining event in p athogenesis. We elucidated genetic, biochemical, and biophysical requiremen ts for assembly of functional type 1 pili, We discovered that the minor pil us protein FimG plays a critical role in nucleating the formation of the ad hesive tip fibrillum, Genetic methods were used to trap pilus subunits duri ng their translocation through the outer membrane usher protein, providing data on the structural interactions that occur between subunit components d uring type 1 pilus formation. Electron microscopic and biochemical analyses of these stepwise assembly intermediates demonstrated that translocation o f pilus subunits occurs linearly through the usher's central channel, with formation of the pilus helix occurring extracellularly, Specialized pilin s ubunits play unique roles both in this multimerization and in the final ult rastructure of the adhesive pilus.