NOVEL OXYGENATIONS OF ARACHIDONIC ACID
Principal Investigator: ALAN BRASH
Affiliation: Vanderbilt University
Abstract: DESCRIPTION: The formation of bioactive mediators, prostaglandins, leukotrienes, and HETEs, is initiated by stereoselective oxygenation of arachidonic acid. The goals of this project are to elucidate the function of novel lipoxygenases in mammalian physiology and to elucidate the enzyme-substrate interactions that distinguish the activities of the different lipoxygenase and cyclooxygenase enzymes. Dr. Brash and his colleagues have discovered several new lipoxygenase cDNAs that are the focus of studies in both the physiological function and the structure-function of lipoxygenase proteins. A primary goal is to characterize a new human 15(S) lipoxygenase and its closely related mouse homologue (a phorbol ester-inducible 8S-lipoxygenase) and to determine their function. These enzymes have been detected in skin, and induction of the murine homologue is known to be associated with inflammation and hyperplasia. The investigators will prepare transgenic mice overexpressing the enzymes in skin to examine the effects on inflammation and carcinogenesis. These human and murine lipoxygenases will also be used in studies of protein structure-function to elucidate determinants of positional specificity in these enzymes, initially using a chimera approach. In relation to the structure-function of lipoxygenases, characterization of the primary structures of several novel enzymes with R stereospecificity will provide valuable new insights on structure-function of these proteins. They also propose to elucidate the enzyme responsible for the increased synthesis of 12R-HETE in human proliferative skin disease: they plan to establish whether this is a R-lipoxygenase or a cytochrome P450 and then to characterize the enzyme involved in the production of this distinctive arachidonic acid metabolite that accumulates in psoriases and other dermatoses. The investigators will also examine the mechanisms of oxygenation of the cyclooxygenases, with comparison and contrast to the R and S lipoxygenases, by analysis of reactions of stereoselectivly-labeled 3H-substrate and the enantioselective reactions with a novel series of substrate analogues in the presence and absence of aspirin. The results of this study will help elucidate the molecular basis for the specificity of oxygenation by different lipoxygenases and cyclooxygenases and characterize an important target for known or potential therapeutic interventions.
Funding Period: 1997-05-01 - 2001-08-31
more information: NIH RePORT
- On the relationships of substrate orientation, hydrogen abstraction, and product stereochemistry in single and double dioxygenations by soybean lipoxygenase-1 and its Ala542Gly mutantGianguido Coffa
Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA
J Biol Chem 280:38756-66. 2005..This provides evidence that both tail-first and carboxylate end-first binding of substrate can be associated with S or R partnerships in product formation in the same active site...
- Convergent oxygenation of arachidonic acid by 5-lipoxygenase and cyclooxygenase-2Claus Schneider
Department of Pharmacology, Division of Clinical Pharmacology, 23rd Avenue South at Pierce, Vanderbilt University Medical School, Nashville, Tennessee 37232, USA
J Am Chem Soc 128:720-1. 2006..The product is a unique diendoperoxide, potentially representing the parent compound of a novel group of lipid mediators...
- Molecular dynamics simulations of arachidonic acid-derived pentadienyl radical intermediate complexes with COX-1 and COX-2: insights into oxygenation regio- and stereoselectivityKristina E Furse
Department of Chemistry, Center for Structural Biology, Vanderbilt University, Nashville, Tennessee 37232, USA
Biochemistry 45:3206-18. 2006..Instead, a combination of oxygen channeling, steric shielding, and selective radical trapping appears to be responsible. This work adds a dynamic perspective to the strong foundation of static structural data available for these enzymes...
- Control of oxygenation in lipoxygenase and cyclooxygenase catalysisClaus Schneider
Department of Pharmacology, Vanderbilt University, Nashville, TN 37232, USA
Chem Biol 14:473-88. 2007..We present four mechanistic models, not mutually exclusive, that could account for the specific reactions of molecular oxygen with a fatty acid in the LOX or COX active site...
- The structure and peroxidase activity of a 33-kDa catalase-related protein from Mycobacterium avium ssp. paratuberculosisSvetlana Pakhomova
Department of Biological Sciences, Louisiana State University, Baton Rouge, Louisiana 70803, USA
Protein Sci 18:2559-68. 2009..Its structural features and the result of the enzyme assays support a role for MAP-2744c and its close homologues in mitigating challenge by a variety of reactive oxygen species...