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Efficient secretion of a folded protein domain by a monomeric bacterial autotransporter
Author(s) -
Skillman Kristen M.,
Barnard Travis J.,
Peterson Janine H.,
Ghirlando Rodolfo,
Bernstein Harris D.
Publication year - 2005
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2005.04885.x
Subject(s) - periplasmic space , biology , bacterial outer membrane , escherichia coli , secretion , biochemistry , c terminus , protein domain , biophysics , microbiology and biotechnology , amino acid , gene
Summary Bacterial autotransporters are proteins that contain a small C‐terminal ‘β domain’ that facilitates translocation of a large N‐terminal ‘passenger domain’ across the outer membrane (OM) by an unknown mechanism. Here we used EspP, an autotransporter produced by Escherichia coli 0157:H7, as a model protein to gain insight into the transport reaction. Initially we found that the passenger domain of a truncated version of EspP (EspPΔ1‐851) was translocated efficiently across the OM. Blue Native polyacrylamide gel electrophoresis, analytical ultracentrifugation and other biochemical methods showed that EspPΔ1‐851 behaves as a compact monomer and strongly suggest that the channel formed by the β domain is too narrow to accommodate folded polypeptides. Surprisingly, we found that a folded protein domain fused to the N‐terminus of EspPΔ1‐851 was efficiently translocated across the OM. Further analysis revealed that the passenger domain of wild‐type EspP also folds at least partially in the periplasm. To reconcile these data, we propose that the EspP β domain functions primarily to target and anchor the protein and that an external factor transports the passenger domain across the OM.