Characterization of the C-terminal domains of intimin-like proteins of enteropathogenic and enterohemorrhagic Escherichia coli, Citrobacter freundii, and Hafnia alvei.
Surface proteins called intimins (Int), which are homologous to the invasin protein (Inv) of Yersinia spp., play a role in inducing brush border damage, termed attachment and effacement, which follows infection by enteropathogenic and enterohemorrhagic Escherichia coli, Citrobacter freundii biotype 4280, and Hafnia alvei. Maltose-binding protein (MBP) fusions containing the C-terminal 280 amino acids of Int-like proteins of strains of enteropathogenic E. coli, enterohemorrhagic E. coli, H. alvei, and C. freundii biotype 4280 and of Yersinia pseudotuberculosis Inv were constructed and purified. The 3' end of the gene for the H. alvei Int-like protein was sequenced and showed homology to corresponding regions of other Int-encoding genes. Binding of MBP-Int-like and MBP-Inv fusion proteins to HEp-2 cells was demonstrated by immunofluorescence microscopy and by fluorescence-activated cell sorting. MBP-Inv induced attachment and spreading of HEp-2 cells to plastic-coated wells, but MBP-Int-like fusion proteins did not. Preincubation of HEp-2 cells with MBP-Inv, but not with MBP-Int-like fusion proteins, inhibited MBP-Inv-induced cell attachment. Fixed staphylococci and fluorescent polymer microspheres coated with both MBP-Int-like and MBP-Inv fusion proteins showed enhanced adhesion to HEp-2 cells. These fusion proteins will facilitate studies of the role of intimin in the pathogenesis of diarrhea associated with members of the family Enterobacteriaeceae that induce attachment and effacement.
Frankel
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Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, United Kingdom.
Candy
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Everest
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Dougan
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D018528:ATP-Binding Cassette Transporters; D018829:Adhesins, Bacterial; D000595:Amino Acid Sequence; D001422:Bacterial Adhesion; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism; D002478:Cells, Cultured; D016971:Citrobacter freundii / Q000472:pathogenicity*; D003967:Diarrhea / Q000209:etiology; D004754:Enterobacter / Q000472:pathogenicity*; D004926:Escherichia coli / Q000472:pathogenicity*; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment
D018528:ATP-Binding Cassette Transporters; D018829:Adhesins, Bacterial; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins; C461599:maltose transport system, E coli; C073819:eaeA protein, E coli