A striking feature of the biology of Haemophilus influenzae is that these organisms can be distinguished from nearly all other facultative, gram-negative bacteria by their absolute requirement for heme. Surprisingly little attention has been paid to understanding how this parasite satisfies this unusual nutritional requirement, especially while growing in its human host. This research project involves a comprehensive approach to this unexplored aspect of H. influenzae molecular biology and disease pathogenesis. The ultimate objective of this research program is the elucidation of the mechanisms used by both typable and nontypable strains of H. influenzae for the acquisition of heme, with particular emphasis on how this pathogen obtains heme from the serum proteins hemopexin and haptoglobin which bind heme and hemoglobin, respectively, in the human body. 1) The newly identified 100kd protein of H. influenzae type b, which binds heme:hemopexin complexes and has been shown to be necessary for utilization of heme bound to hemopexin, will be investigated for its structure-function relationships. In addition, the relationship of this macromolecule to the RTX family of cytotoxins will be determined. 2) The 51kd heme-binding lipoprotein, which is ubiquitous among H. influenzae strains, will be investigated to determine its involvement in transport of heme into the H. influenzae cell. Additional experiments will be performed to determine whether this unusual lipoprotein is a one component of a permease system related to the superfamily of periplasmic binding protein-dependent, ATP-linked transporters. 3) Other H. influenzae cell surface constituents involved in the acquisition of heme from carrier proteins will be identified by the use of molecular genetic techniques, with special emphasis on the nontypable H. influenzae proteins involved in this process. 4) The surface-exposed proteins of nontypable H. influenzae shown to be involved in the acquisition of heme will be evaluated for their potential for vaccine development. This last set of experiments is especially relevant in view of the lack of a vaccine to prevent both upper and lower respiratory tract disease caused by nontypable H. influenzae.
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