Pathogenic bacteria such as Escherichia coli assemble surface structures termed pili, or fimbriae, to mediate binding to host-cell receptors1. Type 1 pili are assembled via the conserved chaperone–usher pathway2,3,4,5. The outer-membrane usher FimD recruits pilus subunits bound by the chaperone FimC via the periplasmic N-terminal domain of the usher. Subunit translocation through the β-barrel channel of the usher occurs at the two C-terminal domains (which we label CTD1 and CTD2) of this protein. How the chaperone–subunit complex bound to the N-terminal domain is handed over to the C-terminal domains, as well as the timing of subunit polymerization into the growing pilus, have previously been unclear. Here we use cryo-electron microscopy to capture a pilus assembly intermediate (FimD–FimC–FimF–FimG–FimH) in a conformation in which FimD is in the process of handing over the chaperone-bound end of the growing pilus to the C-terminal domains. In this structure, FimF has already polymerized with FimG, and the N-terminal domain of FimD swings over to bind CTD2; the N-terminal domain maintains contact with FimC–FimF, while at the same time permitting access to the C-terminal domains. FimD has an intrinsically disordered N-terminal tail that precedes the N-terminal domain. This N-terminal tail folds into a helical motif upon recruiting the FimC-subunit complex, but reorganizes into a loop to bind CTD2 during handover. Because both the N-terminal and C-terminal domains of FimD are bound to the end of the growing pilus, the structure further suggests a mechanism for stabilizing the assembly intermediate to prevent the pilus fibre diffusing away during the incorporation of thousands of subunits.
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The cryo-EM 3D maps of the FimD–tip (FimDCFGH) complex have been deposited at the Electron Microscopy Data Bank database with accession codes EMD-8953 (conformer 1) and EMD-8954 (conformer 2), and their corresponding atomic models were deposited at the PDB with accession codes 6E14 (conformer 1) and 6E15 (conformer 2), respectively.
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Cryo-EM data were collected at the David Van Andel Advanced Cryo-Electron Microscopy Suite at the Van Andel Research Institute. We thank X. Meng for help with data collection. This study was supported by the US National Institutes of Health R01 grants GM062987 (to D.G.T. and H.L.) and GM111742 (to H.L.) and Van Andel Research Institute (to H.L.).
Nature thanks A. Dessen, J. Rubinstein and the other anonymous reviewer(s) for their contribution to the peer review of this work.