Journal
JOURNAL OF MOLECULAR BIOLOGY
Volume 412, Issue 4, Pages 553-567Publisher
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2011.07.035
Keywords
outer membrane; translocation; insertion; passenger; surface
Categories
Funding
- NWO Earth and Life Sciences (The Netherlands)
- NWO (The Netherlands)
- Netherlands Leprosy Relief Foundation
- Aeras Global TB Vaccine Foundation
- Dutch Ministry of Foreign Affairs
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Autotransporters (ATs) of Gram-negative bacteria contain an N-proximal passenger domain that is transported to the extracellular milieu and a C-terminal beta-domain that inserts into the outer membrane (OM) in a beta-barrel conformation. This beta-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 beta-domains show that the beta-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 beta-barrel assembly machinery. These findings questioned a direct involvement of the beta-domain in passenger translocation and suggested that it may only target the passenger to the beta-barrel assembly machinery pore. To address the function of the beta-domain in more detail, we have replaced the beta-domain of the Escherichia coli AT hemoglobin protease by beta-domains originating from other OM proteins. Furthermore, we have modified the diameter of the beta-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 beta-domain that not only functions as a targeting device but also is directly involved in the translocation of the passenger to the cell surface. (C) 2011 Elsevier Ltd. All rights reserved.
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