Secretion of a bacterial virulence factor is driven by the folding of a C-terminal segment.
Autotransporters are bacterial virulence factors consisting of an N-terminal "passenger domain" that is secreted in a C- to-N-terminal direction and a C-terminal "β domain" that resides in the outer membrane (OM). Although passenger domain secretion does not appear to use ATP, the energy source for this reaction is unknown. Here, we show that efficient secretion of the passenger domain of the Escherichia coli O157:H7 autotransporter EspP requires the stable folding of a C-terminal ≈17-kDa passenger domain segment. We found that mutations that perturb the folding of this segment do not affect its translocation across the OM but impair the secretion of the remainder of the passenger domain. Interestingly, an examination of kinetic folding mutants strongly suggested that the ≈17-kDa segment folds in the extracellular space. By mutagenizing the ≈17-kDa segment, we also fortuitously isolated a unique translocation intermediate. Analysis of this intermediate suggests that a heterooligomer that facilitates the membrane integration of OM proteins (the Bam complex) also promotes the surface exposure of the ≈17-kDa segment. Our results provide direct evidence that protein folding can drive translocation and help to clarify the mechanism of autotransporter secretion.
Peterson
Janine H
JH
Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
Tian
Pu
P
Ieva
Raffaele
R
Dautin
Nathalie
N
Bernstein
Harris D
HD
D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D019453:Escherichia coli O157 / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D007700:Kinetics; D009154:Mutation / Q000235:genetics; D017510:Protein Folding; D017434:Protein Structure, Tertiary / Q000235:genetics*; D012697:Serine Endopeptidases / Q000378:metabolism*; D037521:Virulence Factors / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; D037521:Virulence Factors; C107189:EspP protein, E coli; D012697:Serine Endopeptidases