title: Disulfide bond-mediated passenger domain stalling as a structural probe of autotransporter outer membrane secretion in vivo.
abstract: Autotransporters (ATs) are the largest class of extracellular virulence proteins secreted by Gram-negative pathogenic bacteria, but the details of their outer membrane (OM) secretion mechanism remain unclear. Recently, a novel strategy has been developed to study OM secretion of AT proteins by introducing pairs of cysteine (Cys) residues into the central passenger domain sequence. Upon oxidation in the periplasm, these Cys residues form a long loop that stalls AT OM secretion. This Cys-loop stalling technique has been used to investigate such questions as the directionality of AT OM secretion and the extent of AT passenger domain folding during secretion. Here, we will describe how to use the Cys-loop approach to produce disulfide-bonded, stalled AT OM secretion intermediates, and how these stalled "snapshots" can be used to investigate structural aspects of the AT OM secretion mechanism.
authors.affiliation: Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, Indiana, USA.
authors.forename: Jonathan P
authors.initials: JP
authors.lastname: Renn
authors.forename: Patricia L
authors.initials: PL
authors.lastname: Clark
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry; D006090:Gram-Negative Bacteria / Q000737:chemistry* / Q000472:pathogenicity; D008958:Models, Molecular; D017510:Protein Folding
chemical_list: D001425:Bacterial Outer Membrane Proteins; D003545:Cysteine