title: The periplasmic disulfide oxidoreductase DsbA contributes to Haemophilus influenzae pathogenesis.
abstract: Haemophilus influenzae is an obligate human pathogen that persistently colonizes the nasopharynx and causes disease when it invades the bloodstream, lungs, or middle ear. Proteins that mediate critical interactions with the host during invasive disease are likely to be secreted. Many secreted proteins require addition of disulfide bonds by the DsbA disulfide oxidoreductase for activity or stability. In this study, we evaluated the role in H. influenzae pathogenesis of DsbA, as well as HbpA, a substrate of DsbA. Mutants of H. influenzae Rd and type b strain Eagan having nonpolar deletions of dsbA were attenuated for bacteremia in animal models, and complemented strains exhibited virulence equivalent to that of the parental strains. Comparison of predicted secreted proteins in H. influenzae to known DsbA substrates in other species revealed several proteins that could contribute to the role of dsbA in virulence. One candidate, the heme transport protein, HbpA, was examined because of the importance of exogenous heme for aerobic growth of H. influenzae. The presence of a dsbA-dependent disulfide bond in HbpA was verified by an alkylation protection assay, and HbpA was less abundant in a dsbA mutant. The hbpA mutant exhibited reduced bacteremia in the mouse model, and complementation restored its in vivo phenotype to that of the parental strain. These results indicate that dsbA is required in vivo and that HbpA and additional DsbA-dependent factors are likely to participate in H. influenzae pathogenesis.
authors.affiliation: Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, 55 Lake Ave., N., S6-242, Worcester, MA 01655, USA.
authors.forename: Charles V
authors.initials: CV
authors.lastname: Rosadini
authors.forename: Sandy M S
authors.initials: SM
authors.lastname: Wong
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Akerley
mesh_terms: D000818:Animals; D016470:Bacteremia / Q000382:microbiology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D005260:Female; D017353:Gene Deletion; D006192:Haemophilus Infections / Q000382:microbiology; D006193:Haemophilus influenzae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000472:pathogenicity*; D006418:Heme / Q000378:metabolism; D008074:Lipoproteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D019897:Periplasm / Q000201:enzymology*; D019704:Protein Disulfide-Isomerases / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015996:Survival Rate; D014774:Virulence
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; C097952:HbpA protein, Haemophilus influenzae; D008074:Lipoproteins; D006418:Heme; D019704:Protein Disulfide-Isomerases