The coiled-coil domain of EspA is essential for the assembly of the type III secretion translocon on the surface of enteropathogenic Escherichia coli.
Enteropathogenic E. coli (EPEC) utilize a type III secretion system to deliver virulence-associated effector proteins to the host cell. Four proteins, EspA, EspB, EspD, and Tir, which are integral to the formation of characteristic "attaching and effacing" (A/E) intestinal lesions, are known to be exported via the EPEC type III secretion system. Recent work demonstrated that EspA is a major component of a filamentous structure, elaborated on the surface of EPEC, which is required for translocation of EspB and Tir. The carboxyl terminus of EspA is predicted to comprise an alpha-helical region, which demonstrates heptad periodicity whereby positions a and d in the heptad repeat unit abcdefg are occupied by hydrophobic residues, indicating a propensity for coiled-coil interactions. Here we demonstrate multimeric EspA isoforms in EPEC culture supernatants and EspA:EspA interaction on solid phase. Non-conservative amino acid substitution of specific EspA heptad residues generated EPEC mutants defective in filament assembly but which retained the ability to induce A/E lesions; additional mutation totally abolished EspA filament assembly and A/E lesion formation. These results demonstrate a similarity to flagellar biosynthesis and indicate that the coiled-coil domain of EspA is required for assembly of the EspA filament-associated type III secretion translocon.
Delahay
R M
RM
Department of Biochemistry, Imperial College of Science, Technology and Medicine, London SW7 2AZ, United Kingdom.
Knutton
S
S
Shaw
R K
RK
Hartland
E L
EL
Pallen
M J
MJ
Frankel
G
G
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D002462:Cell Membrane / Q000502:physiology*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics* / Q000472:pathogenicity; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry; D015398:Signal Transduction; D012984:Software; D014774:Virulence
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C102051:EspA protein, E coli; D011993:Recombinant Fusion Proteins