title: Functional comparison of serine protease autotransporters of enterobacteriaceae.
abstract: The plasmid-encoded toxin (Pet) of enteroaggregative Escherichia coli (EAEC) belongs to a family of high-molecular-weight serine protease autotransporters of Enterobacteriaceae (SPATEs) which also includes Pic from EAEC and Shigella flexneri, EspC from enteropathogenic E. coli, EspP from enterohemorrhagic E. coli, Sat from uropathogenic E. coli, Tsh from avian pathogenic E. coli, and SepA from S. flexneri. Phylogenetic analysis shows the SPATE proteins to represent a distinct subfamily of autotransporters with amino acid identities ranging from 35 to 55%, providing a powerful resource to direct structure-function studies. In this study, we show that these related proteins are proteases with divergent substrate specificities, suggesting different functions. The cleavage profile of oligopeptides was found to be unique for each SPATE protein. The SPATEs showed proteolytic activity for several substrates, namely mucin, pepsin, human coagulation factor V, and erythroid spectrin. The cleavage of spectrin has been hypothesized as the mechanism through which Pet induces cytopathic effects. However, whereas Pet, Sat, and EspC cleaved spectrin, only Pet and Sat elicited cytopathic effects; the remaining SPATEs did not cause any morphological changes to HEp-2 cell monolayers. EspC and Pet exhibited acid-dissociable binding to HEp-2 cells. However, Pet was more efficient at entering HEp-2 cells, suggesting a basis for the different abilities of these two proteases to damage cells. Our data suggest that, despite the homologies observed among these proteins, the SPATEs have different pathogenetic functions only partly dependent on their substrate specificities.
authors.affiliation: Center for Vaccine Development, University of Maryland School of Medicine, Baltimore 21201, USA.
authors.forename: Pinaki R
authors.initials: PR
authors.lastname: Dutta
authors.forename: Renato
authors.initials: R
authors.lastname: Cappello
authors.forename: Fernando
authors.initials: F
authors.lastname: Navarro-García
authors.forename: James P
authors.initials: JP
authors.lastname: Nataro
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004755:Enterobacteriaceae / Q000201:enzymology* / Q000472:pathogenicity; D004847:Epithelial Cells / Q000473:pathology; D006801:Humans; D010802:Phylogeny; D010957:Plasmids / Q000235:genetics; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D012697:Serine Endopeptidases