Structure and Assembly of a Trans-Periplasmic Channel for Type IV Pili in Neisseria meningitidis

Type IV pili are polymeric fibers which protrude from the cell surface and play a critical role in adhesion and invasion by pathogenic bacteria. The secretion of pili across the periplasm and outer membrane is mediated by a specialized secretin protein, PilQ, but the way in which this large channel is formed is unknown. Using NMR, we derived the structures of the periplasmic domains from N. meningitidis PilQ: the N-terminus is shown to consist of two beta-domains, which are unique to the type IV pilus-dependent secretins. The structure of the second beta-domain revealed an eight-stranded beta-sandwich structure which is a novel variant of the HSP20-like fold. The central part of PilQ consists of two alpha/beta fold domains: the structure of the first of these is similar to domains from other secretins, but with an additional alpha-helix which links it to the second alpha/beta domain. We also determined the structure of the entire PilQ dodecamer by cryoelectron microscopy: it forms a cage-like structure, enclosing a cavity which is approximately 55 angstrom in internal diameter at its largest extent. Specific regions were identified in the density map which corresponded to the individual PilQ domains: this allowed us to dock them into the cryoelectron microscopy density map, and hence reconstruct the entire PilQ assembly which spans the periplasm. We also show that the C-terminal domain from the lipoprotein PilP, which is essential for pilus assembly, binds specifically to the first alpha/beta domain in PilQ and use NMR chemical shift mapping to generate a model for the PilP: PilQ complex. We conclude that passage of the pilus fiber requires disassembly of both the membrane-spanning and the b-domain regions in PilQ, and that PilP plays an important role in stabilising the PilQ assembly during secretion, through its anchorage in the inner membrane.