Folding of a bacterial integral outer membrane protein is initiated in the periplasm.

ABSTRACT

The Bam complex promotes the insertion of ß-barrel proteins into the bacterial outer membrane, but it is unclear whether it threads ß-strands into the lipid bilayer in a stepwise fashion or catalyzes the insertion of pre-folded substrates. Here, to distinguish between these two possibilities, we analyze the biogenesis of UpaG, a trimeric autotransporter adhesin (TAA). TAAs consist of three identical subunits that together form a single ß-barrel domain and an extracellular coiled-coil ("passenger") domain. Using site-specific photocrosslinking to obtain spatial and temporal insights into UpaG assembly, we show that UpaG ß-barrel segments fold into a trimeric structure in the periplasm that persists until the termination of passenger-domain translocation. In addition to obtaining evidence that at least some ß-barrel proteins begin to fold before they interact with the Bam complex, we identify several discrete steps in the assembly of a poorly characterized class of virulence factors.