The objective of this proposal is to elucidate the protein-ligand interactions and structure/function relationships in three new bacterial hemoglobins (Hb) and two mammalian prostaglandin H synthases (PGHS-1 and PGHS-2). The two bacterial hemoglobins from Mycobacterium tuberculosis (HbN and HbO) belong to a newly discovered truncated hemoglobin family, which are characterized by a novel two-over-two alpha-helical sandwich motif, the absence of the A- helix and the presence of an extended loop substituting for most of the F-helix. The physiological functions of HbN and HbO are not established but because O2 delivery in unicellular organisms is a diffusion-controlled process, functions other than oxygen transport have been put forth. The bacterial hemoglobin from E. coli (Hmp) is a flavohemoglobin consisting of a heme-containing globin-like domain and a FAD-containing reductase domain. It is believed that the function of Hmp is to detoxify NO and other reactive nitrogen species. The structural properties of the three bacterial hemoglobins will be fully characterized. Based on our preliminary resonance Raman studies, we postulate that the heme pockets of these bacterial hemoglobins are tailored to perform chemistry, such as oxygen activation, and that they may share structural and functional similarities to peroxidases. This hypothesis will be tested by studies of the reactions of these hemoglobins with NO, hydrogen peroxide and peroxynitrite. The possible role of these hemoglobins in protecting the microorganisms against attack by reactive nitrogen intermediates will be explored by monitoring NO and O2 consumption in wild-type and hemoglobin knock-out cells. Related reactions will be studied in the peroxidase sites of two PGHS isoforms, which play an essential role in the synthesis of prostaglandins. Preliminary data suggests that the proximal bond that coordinates the heme to the polypeptide is quite weak in PGHS's, which is very unusual for peroxidases. Experiments are proposed to test the functional consequences of this finding. These hemeprotein systems provide an excellent model for investigating fundamental structural properties that underlie biological reactivity.
|Effective start/end date||12/1/01 → 5/31/06|
- Infectious Diseases
- Structural Biology
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