Mapping the epithelial-cell-binding domain of the Aggregatibacter actinomycetemcomitans autotransporter adhesin Aae

Daniel H. Fine, Jeffrey B. Kaplan, David Furgang, Maribasappa Karched, Kabilan Velliyagounder, Gang Yue

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

The Gram-negative periodontopathogen Aggregatibacter actinomycetemcomitans (Aa) binds selectively to buccal epithelial cells (BECs) of human and Old World primates by means of the outer-membrane autotransporter protein Aae. We speculated that the exposed N-terminal portion of the passenger domain of Aae would mediate binding to BECs. By using a series of plasmids that express full-length or truncated Aae proteins in Escherichia coli, we found that the BEC-binding domain of Aae was located in the N-terminal surface-exposed region of the protein, specifically in the region spanning amino acids 201-284 just upstream of the repeat region within the passenger domain. Peptides corresponding to amino acids 201-221, 222-238 and 201-240 were synthesized and tested for their ability to reduce Aae-mediated binding to BECs based on results obtained with truncated Aae proteins expressed in E. coli. BEC-binding of E. coli expressing Aae was reduced by as much as 50% by pre-treatment of BECs with a 40-mer peptide (201-240; P40). Aae was also shown to mediate binding to cultured human epithelial keratinocytes (TW2.6), OBA9 and TERT, and endothelial (HUVEC) cells. Pre-treatment of epithelial cells with P40 resulted in a dose-dependent reduction in binding and reduced the binding of both full-length and truncated Aae proteins expressed in E. coli, as well as Aae expressed in Aa. Fluorescently labelled P40 peptides reacted in a dose-dependent manner with BEC receptors. We propose that these proof-of-principle experiments demonstrate that peptides can be designed to interfere with Aa binding mediated by host-cell receptors specific for Aae adhesins.

Original languageEnglish (US)
Pages (from-to)3412-3420
Number of pages9
JournalMicrobiology
Volume156
Issue number11
DOIs
StatePublished - Nov 2010

All Science Journal Classification (ASJC) codes

  • Microbiology

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