Structural insight into the assembly of the type II secretion system pilotin-secretin complex from enterotoxigenic Escherichia coli.

Secretin is a large outer-membrane channel found in secretion systems of Gram-negative bacteria, facilitating the last step for transfer of proteins into the extracellular environment. In the type II secretion system, a lipoprotein called pilotin is essential to bind and target its corresponding secretin to the outer membrane. However, there is only limited structural information available about the interaction and assembly of the pilotin-secretin complex. Here we report the first near-atomic-resolution structure of a full-length Vibrio-type pilotin-secretin (AspS-GspD) complex from enterotoxigenic Escherichia coli by cryo-electron microscopy, which reveals the detailed assembly mode of the full-length pilotin-secretin complex. The AspS subunits attach to the secretin channel surface with a 15:15 stoichiometric ratio to GspD subunits, and insert their amino terminus into the outer membrane. The AspS subunits interact with all three secondary structural elements of the S domain of GspD, including strong interaction with the carboxy-terminal α-helix and weak interactions with another two elements, an α-helix and a loop. These structural and biochemical details provide a deeper insight to pilotin-secretin interaction and their assembly mode.