Abstract

Autotransporters are virulence factors produced by Gram-negative bacteria. They consist of two domains, an N-terminal 'passenger' domain and a C-terminal β-domain. β-domains form β-barrel structures in the outer membrane while passenger domains are translocated into the extracellular space. In some autotransporters, the two domains are separated by proteolytic cleavage. Using X-ray crystallography, we solved the 2.7-|[Aring]| structure of the post-cleavage state of the β-domain of EspP, an autotransporter produced by Escherichia coli strain O157:H7. The structure consists of a 12-stranded β-barrel with the passenger domain–β-domain cleavage junction located inside the barrel pore, approximately midway between the extracellular and periplasmic surfaces of the outer membrane. The structure reveals an unprecedented intra-barrel cleavage mechanism and suggests that two conformational changes occur in the β-domain after cleavage, one conferring increased stability on the β-domain and another restricting access to the barrel pore.