This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Lipid metabolism is a major human health concern, with much attention focused on inflammation and lipid homeostatis of lipids linked to low and high density lipoprotein (LDL, HDL). Many proteins that are associated with LDL and HDL particles play a critical role in these lipid pathways. The plasma form of platelet activating factor acetylhydrolase (pPAFAH) functions on the surface of LDL particles by reducing levels of the signaling molecule platelet activating factor (PAF) as a general anti-inflammatory scavenger, and is linked to anaphylactic shock, asthma and allergic reactions. As a LDL-associated protein with no known homologues, pPAFAH is a worthy structural target. In addition to its role to reduce PAF levels, it has also been implicated in hydrolytic activities of other pro-inflammatory agents, such as oxidized lipids of LDL particles. A molecular understanding of the physiological reactions catalyzed by pPAFAH requires structural models of the enzyme interacting with its substrates PAF and oxidized phospholipids. We will elucidate the relationship between structure and interfacial enzyme function for pPAFAH via 4 specific aims: (i) The high-resolution crystal structure of pPAFAH will be solved. The use of detergents and amphiphilic molecules will be explored to provide higher resolution structures, as well as functionally more relevant structures. (ii) Inhibitors of pPAFAH will be explored and developed via structural and kinetic characterization to elucidate in vivo physiological functions. (iii) Combining structure with interfacial kinetic studies, we will further elucidate the physiological roles of pPAFAH. Insights from inhibition and structural studies will help develop an atomic resolution mechanism of the enzyme. Enzyme-ligand complexes will be pursued that further elucidate the enzyme mechanism. (iv) The physiologically relevant reaction of pPAFAH with organophosphate (OP) compounds will be explored, including the possible use of the LDL-associated enzyme as a bioscavenger target of OP neurotoxins.
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