Structure and characterization of AgfB from Salmonella enteritidis thin aggregative fimbriae

The agfBAC operon of Salmonella enteritidis encodes thin aggregative fimbri ae, fibrous, polymeric structures primarily composed of AgfA fimbrins. Alth ough uncharacterized, AgfB shows a 51% overall amino acid sequence similari ty to AgfA. Using AgfB epitope-specific antiserum, AgfB was detected as a m inor component of whole, purified fimbriae. Like AgfA, AgfB was released fr om purified fimbriae by >70% formic acid, whereupon both AgfA-AgfA and AgfA -AgfB dimers as well as monomers were detected. This suggested that AgfB ma y form specific, highly stable, structural associations with AgfA in native fimbrial filaments, associations that were weakened in structurally unstab le fibers derived from AgfA chimeric fimbrial mutants. Detailed sequence co mparisons between AgfA and AgfB showed that AgfB harbored a similar fivefol d repeated sequence pattern (x(6)QxGx(2)NxAx(3)Q), and contained structural motifs similar to the parallel beta helix model proposed for AgfA. Molecul ar modeling of AgfB revealed a 3D structure remarkably similar to that of A gfA, the structures differing principally in the surface disposition of non -conserved, basic, acidic and non-polar residues. Thus AgfB is a fimbrin-li ke structural homologue of AgfA and an integral, minor component of native thin aggregative fimbrial fibers. AgfB from an agfA deletion strain was det ected as a non-fimbrial, SDS-insoluble form in the supernatant and was puri fied. AgfA from an agfB deletion strain was found in both SDS-soluble and i nsoluble, non-fimbrial forms. No AgfA-AgfA dimers were detected in the abse nce of AgfB. Fimbriae formation by intercellular complementation between ag fB and agfA deletion strains could not be shown under a variety of conditio ns, indicating that AgfA and AgfB are not freely diffusible in S. enteritid is. This has important implications on the current assembly hypothesis for thin aggregative fimbriae. (C) 2001 Academic Press.