Haemophilus influenzae is a common cause of localized respiratory tract disease, including otitis media, sinusitis, bronchitis, and pneumonia. Less commonly, this organism causes serious systemic disease, such as meningitis, septicemia, and endocarditis. The initial step in the pathogenesis of H. influenzae disease involves colonization of the upper respiratory mucosa. We have identified a non-pilus adhesin called Hia, which is present in nontypable (nonencapsulated) H. influenzae and promotes high-affinity attachment to human epithelium. In addition, we have identified a homolog of Hia called Hsf, which is present in nearly all encapsulated strains of H. influenzae and also mediates high-affinity adherence to epithelial cells. Both Hia and Hsf are members of the expanding family of auto-transporter proteins, characterized by an N-terminal signal peptide, an internal passenger domain, and a C-terminal trans-locator domain. In contrast to classic auto-transporters, Hia and Hsf have a translocator domain that is unusually short and forms a trimer. For both Hia and Hsf, the passenger domain has two distinct binding regions that interact with the same host cell receptor, although with differing affinities. We have crystallized a polypeptide containing the Hia primary binding domain and find a densely packed trimer with three-fold symmetry and three identical major grooves, acidic pockets, and globular domains.
In Aim 1 of this proposal, we will define the surfaces of the Hia and Hsf binding domains involved in interaction with host cells. In addition, we will explore whether trimer formation is required for adhesive activity.
In Aim 2, we will characterize the structure and mechanism of action of the Hia and Hsf translocator domain.
In Aim 3, we will identify the Hia/Hsf host cell receptor and elucidate the host cell response to Hia and Hsf-mediated adherence. From a practical perspective, the results of these experiments may facilitate efforts to develop a vaccine protective against non-type b H. influenzae and suggest targets for novel antimicrobials with activity against a broad range of gram-negative bacteria. More generally, they may provide fundamental insights into host-pathogen relations, the autotransporter family of proteins, and protein secretion.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Bacteriology and Mycology Subcommittee 2 (BM)
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Khambaty, Farukh M
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Duke University
Schools of Medicine
United States
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Radin, Jana N; Grass, Susan A; Meng, Guoyu et al. (2009) Structural basis for the differential binding affinities of the HsfBD1 and HsfBD2 domains in the Haemophilus influenzae Hsf adhesin. J Bacteriol 191:5068-75
Sheets, Amanda J; Grass, Susan A; Miller, Sara E et al. (2008) Identification of a novel trimeric autotransporter adhesin in the cryptic genospecies of Haemophilus. J Bacteriol 190:4313-20
Meng, Guoyu; St Geme 3rd, Joseph W; Waksman, Gabriel (2008) Repetitive architecture of the Haemophilus influenzae Hia trimeric autotransporter. J Mol Biol 384:824-36
Cotter, Shane E; Surana, Neeraj K; Grass, Susan et al. (2006) Trimeric autotransporters require trimerization of the passenger domain for stability and adhesive activity. J Bacteriol 188:5400-7
Meng, Guoyu; Surana, Neeraj K; St Geme 3rd, Joseph W et al. (2006) Structure of the outer membrane translocator domain of the Haemophilus influenzae Hia trimeric autotransporter. EMBO J 25:2297-304
Sukupolvi-Petty, Soila; Grass, Susan; St Geme 3rd, Joseph W (2006) The Haemophilus influenzae Type b hcsA and hcsB gene products facilitate transport of capsular polysaccharide across the outer membrane and are essential for virulence. J Bacteriol 188:3870-7
Cotter, Shane E; Surana, Neeraj K; St Geme 3rd, Joseph W (2005) Trimeric autotransporters: a distinct subfamily of autotransporter proteins. Trends Microbiol 13:199-205
Cotter, Shane E; Yeo, Hye-Jeong; Juehne, Twyla et al. (2005) Architecture and adhesive activity of the Haemophilus influenzae Hsf adhesin. J Bacteriol 187:4656-64
Surana, Neeraj K; Cutter, David; Barenkamp, Stephen J et al. (2004) The Haemophilus influenzae Hia autotransporter contains an unusually short trimeric translocator domain. J Biol Chem 279:14679-85
Surana, Neeraj K; Grass, Susan; Hardy, Gail G et al. (2004) Evidence for conservation of architecture and physical properties of Omp85-like proteins throughout evolution. Proc Natl Acad Sci U S A 101:14497-502

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