G. Frankel et al., MOLECULAR CHARACTERIZATION OF A CARBOXY-TERMINAL EUKARYOTIC-CELL-BINDING DOMAIN OF INTIMIN FROM ENTEROPATHOGENIC ESCHERICHIA-COLI, Infection and immunity, 63(11), 1995, pp. 4323-4328
A eukaryotic cell-binding domain from the intimin (Int) polypeptide of
enteropathogenic Escherichia coli O127 (EPEC) was investigated, Deriv
atives of the carboxy-terminal 280-amino-acid domains of Int (Int(EPEC
280)) and the Int homolog invasin (Inv) from Yersinia pseudotuberculos
is (Inv(YP280)) were fused to the E. call maltose-binding protein (MBP
), expressed, and purified. The smallest MBP-Int(EPEC) fusion protein
that efficiently mediated binding to HEp-2 cells, monitored by using p
urified fusion proteins in fluorescence activated cell sorter analysis
or by using fluorescent Covaspheres coated with purified fusions, con
tained the carboxy-terminal 150 amino acids of Int. Replacement of Cys
-937 with Ser (Int(EPEC280CS)) destroyed the cell-binding activity of
Int(EPEC280). Covaspheres coated with MBP-Int(EPEC280) were associated
with HEp-2 cell microvilli but failed to induce actin accumulation un
derneath bound particles or cell spreading on coated plastic surfaces,
MBP-Int(EPEC280), but not MBP, MBP-Int(EPEC280CS), or MBP-Inv(YP280),
inhibited EPEC entry into HEp-2 cells.