Haemophilus influenzae is 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, endocarditis, and septicemia. The initial step in the pathogenesis of H. influenzae disease involves colonization of the upper respiratory mucosa. We have identified a high-molecular-weight protein called Hia, which is present in nontypable (nonencapsulated) H. influenzae and promotes attachment to human epithelium. In addition, we have identified a homolog of Hia called Hsf, which is univerally present among encapsulated H. influenzae and also mediates in vitro adherence. Interestingly, Hia has a predicted molecular mass of approximately 114 kDa and is not detectable by coventional transmission electron microscopy, while Hsf has a predicted molecular mass of approximately 245 kDa and is associated with the presence of short, thin surface fibrils visible by negative staining electron microscopy. Based on our in vitro results, we speculate that Hia and Hsf are important colonization factors. In the present proposal, we plan to characterize the pathway by which Hia and Hsf are localized on the surface of the organism. In particular, we will define the structural features of these proteins that direct them to the periplasm and facilitate their translocation across the outer membrane. We will dissect the influence of an unusual N-terminal extremity, a C-terminal domain predicted to form a beta-barrel, and a putative ATP-binding motif. In additional studies, we will examine the architecture of Hia and will investigate the relationship between structure and adhesive activity, focusing in particular on the role of a predicted coiled coil motif. We will also determine whether Hia and Hsf function interchangeably in an encapsulated strain. 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 the biogenesis of non-pilus adhesins and the nature of the host-microbial relationship.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
1R01AI044167-01
Application #
2738138
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Program Officer
Klein, David L
Project Start
1998-12-15
Project End
2003-11-30
Budget Start
1998-12-15
Budget End
1999-11-30
Support Year
1
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Washington University
Department
Pediatrics
Type
Schools of Medicine
DUNS #
062761671
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
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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