Mechanistic link between β barrel assembly and the initiation of autotransporter secretion.
ABSTRACT Autotransporters are bacterial virulence factors that contain an N-terminal extracellular ("passenger") domain and a C-terminal β barrel ("β") domain that anchors the protein to the outer membrane. The β domain is required for passenger domain secretion, but its exact role in autotransporter biogenesis is unclear. Here we describe insights into the function of the β domain that emerged from an analysis of mutations in the Escherichia coli O157:H7 autotransporter EspP. We found that the G1066A and G1081D mutations slightly distort the structure of the β domain and delay the initiation of passenger domain translocation. Site-specific photocrosslinking experiments revealed that the mutations slow the insertion of the β domain into the outer membrane, but do not delay the binding of the β domain to the factor that mediates the insertion reaction (the Bam complex). Our results demonstrate that the β domain does not simply target the passenger domain to the outer membrane, but promotes translocation when it reaches a specific stage of assembly. Furthermore, our results provide evidence that the Bam complex catalyzes the membrane integration of β barrel proteins in a multistep process that can be perturbed by minor structural defects in client proteins.
- SourceAvailable from: Simran Jeet Kaur[Show abstract] [Hide abstract]
ABSTRACT: The genus Rickettsia (Alphaproteobacteria; Rickettsiales; Rickettsiaceae) is comprised of obligate intracellular parasites, with virulent species of interest both as causes of emerging infectious diseases and for their potential deployment as bioterrorism agents. Currently there are no effective commercially available vaccines, with treatment limited primarily to tetracycline antibiotics, though others (e.g., josamycin, ciprofloxacin, chloramphenicol and azithromycin) are also effective. Much of the recent research geared towards understanding mechanisms underlying rickettsial pathogenicity has centered on characterization of secreted proteins that directly engage eukaryotic cells. Herein, we review all aspects of the Rickettsia secretome, including six secretion systems, 19 characterized secretory proteins, and potential moonlighting proteins identified on surfaces of multiple Rickettsia species. Employing bioinformatics and phylogenomics, we present novel structural and functional insight on each secretion system. Unexpectedly, our investigation revealed that the majority of characterized secretory proteins have not been assigned to their cognate secretion pathways. Furthermore, for most secretion pathways, the requisite signal sequences mediating translocation are poorly understood. As a blueprint for all known routes of protein translocation into host cells, this resource will assist research aimed at uniting characterized secreted proteins with their apposite secretion pathways. Furthermore, our work will help in the identification of novel secreted proteins involved in rickettsial “life on the inside”.This article is protected by copyright. All rights reserved.FEMS microbiology reviews 08/2014; 39(1). DOI:10.1111/1574-6976.12084 · 13.81 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Abstract Pathogenic Gram-negative bacteria have evolved several secretion mechanisms to translocate adhesins, enzymes, toxins and other virulence factors across the inner and outer membrane. Currently, eight different secretion systems, type I - type VIII (T1SS - T8SS) plus the chaperone-usher (CU) pathway have been identified, which act in one-step or two-step mechanisms to traverse both membrane barriers. The type V secretion system (T5SS) is dependent first on the Sec translocon within the inner membrane. The periplasmic intermediates are then secreted through aqueous pores formed by β-barrels in the outer membrane. Until now, transport across the outer membrane has not been understood on a molecular level. With respect to special characteristics revealed by crystal structure analysis, bioinformatic and biochemical data, five subgroups of T5SS were defined. Here, we compare the transport moieties of members of four subgroups based on X-ray crystal structures. For the fifth subgroup, which was identified only recently no structures have so far been reported. We also discuss different models for the translocation process across the outer membrane with respect to recent findings.Biological Chemistry 08/2013; 394(11). DOI:10.1515/hsz-2013-0162 · 2.69 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: TamA is an Escherichia coli Omp85 protein involved in autotransporter biogenesis. It comprises a 16-stranded transmembrane β-barrel and three POTRA domains. The 2.3-Å crystal structure reveals that the TamA barrel is closed at the extracellular face by a conserved lid loop. The C-terminal β-strand of the barrel forms an unusual inward kink, which weakens the lateral barrel wall and creates a gate for substrate access to the lipid bilayer.Nature Structural & Molecular Biology 09/2013; 20(11). DOI:10.1038/nsmb.2689 · 11.63 Impact Factor