PLEIOTROPIC DEFENSES--THE HEMOPEXIN SYSTEM

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DESCRIPTION (taken from the applicant's abstract) The term "pleiotropic defense" is used here to refer to biological systems that act to defend the organism against more than one deleterious agent or condition. The properties of the hemopexin system, which carries out receptor-mediated heme transport by endocytosis of hemopexin and its plasma membrane receptor, make it a paradigm for such protective mechanisms, and these two proteins are the foci of this R03 research proposal. The hemopexin system plays several distinct yet connected roles as part of the body's defense mechanisms, acting as an anti-oxidant in trauma, hemolysis and inflammation, a sequester of heme from pathogens, a conserver of the cell nutrient iron, and an inducer of protective proteins like ferritin and metallothionein. The specific aims for the two years of requested support are: (1) to isolate by affinity chromatography sufficient human hemopexin receptor to allow: (a) initial characterization of the receptor, i.e., molecular weight, subunit composition, and glycosylation, (b) production of specific high affinity antibodies to the receptor, using the services of a commercial facility, and (c) cloning and sequencing the cDNA for the human hemopexin receptor, and (2) to continue to characterize the structural determinants of the hemopexin-hemopexin receptor interaction. To accomplish these objectives, standard yet robust techniques that are already established in this laboratory will be employed, including affinity chromatography for receptor isolation; whole cell, ligand blot and soluble receptor binding assays; cDNA cloning and sequencing and site-directed mutagenesis. We will also obtain tools for future research on the structure and regulation of the receptor and basic information on the interaction of hemopexin with its receptor. Such information is needed to develop therapeutic measures for conditions like re-perfusion injury. Information on the mammalian receptor will also complement research on the human pathogen bacterial hemopexin receptors and provide insight needed to design means to selectively inhibit bacterial, but not host, hemopexin receptors.