Dissecting the Escherichia coli periplasmic chaperone network using differential proteomics

β‐Barrel proteins, or outer membrane proteins ( OMP s), perform many essential functions in G ram‐negative bacteria, but questions remain about the mechanism by which they are assembled into the outer membrane ( OM ). In E scherichia coli , β‐barrels are escorted across the periplasm by chaperones, most notably S ur A and S kp. However, the contributions of these two chaperones to the assembly of the OM proteome remained unclear. We used differential proteomics to determine how the elimination of S kp and S ur A affects the assembly of many OMP s. We have shown that removal of S kp has no impact on the levels of the 63 identified OM proteins. However, depletion of S ur A in the skp strain has a marked impact on the OM proteome, diminishing the levels of almost all β‐barrel proteins. Our results are consistent with a model in which SurA plays a primary chaperone role in E . coli . Furthermore, they suggest that while no OMP s prefer the S kp chaperone pathway in wild‐type cells, most can use S kp efficiently when S ur A is absent. Our data, which provide a unique glimpse into the protein content of the nonviable sur A skp mutant, clarify the roles of the periplasmic chaperones in E . coli .