title: Autotransporter β-domains have a specific function in protein secretion beyond outer-membrane targeting.
abstract: Autotransporters (ATs) of Gram-negative bacteria contain an N-proximal passenger domain that is transported to the extracellular milieu and a C-terminal β-domain that inserts into the outer membrane (OM) in a β-barrel conformation. This β-domain facilitates translocation of the passenger domain across the OM and has long been considered to be the translocation pore. However, available crystal structures of β-domains show that the β-barrel pore is too narrow for the observed transport of folded elements within the passenger domains. ATs have recently been shown to interact with the β-barrel assembly machinery. These findings questioned a direct involvement of the β-domain in passenger translocation and suggested that it may only target the passenger to the β-barrel assembly machinery pore. To address the function of the β-domain in more detail, we have replaced the β-domain of the Escherichia coli AT hemoglobin protease by β-domains originating from other OM proteins. Furthermore, we have modified the diameter of the β-domain pore. The mutant proteins were analyzed for their capacity to insert into the OM and for surface display of the passenger. Our results show that efficient passenger secretion requires a specific β-domain that not only functions as a targeting device but also is directly involved in the translocation of the passenger to the cell surface.
authors.affiliation: Section of Molecular Microbiology, Department of Molecular Cell Biology, VU University, 1081 HV Amsterdam, The Netherlands. ana.sauri@falw.vu.nl
authors.forename: Ana
authors.initials: A
authors.lastname: Saurí
authors.forename: Nadia
authors.initials: N
authors.lastname: Oreshkova
authors.forename: Zora
authors.initials: Z
authors.lastname: Soprova
authors.forename: Wouter S P
authors.initials: WS
authors.lastname: Jong
authors.forename: Musa
authors.initials: M
authors.lastname: Sani
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Peters
authors.forename: Joen
authors.initials: J
authors.lastname: Luirink
authors.forename: Peter
authors.initials: P
authors.lastname: van Ulsen
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D008954:Models, Biological; D008958:Models, Molecular; D030781:Organisms, Genetically Modified; D017510:Protein Folding; D017434:Protein Structure, Tertiary / Q000235:genetics / Q000502:physiology*; D021381:Protein Transport / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins