Nitric oxide synthase (NOS) converts arginine to citrulline and the radical diatomic, nitric oxide. Nitric oxide is now well established as a critical messenger molecule in the neural, endothelial and immune systems for both homeostasis and function. High levels of nitric oxide, however, have been also proven to be responsible for toxic shock, cardiac disease and sever brain damage. Inhibition of this enzyme has therefore become of primary importance. Nitric oxide is synthesized in various tissues by select isoforms and ubiquitous nature of nitric oxide production has severe consequences for NOS inhibition since unreasonable side-effects have already been shown to exist during nonselective inhibition. Although the isoforms are highly homologous, we have shown that critical differences between the enzymes do exist and that the active sites of the enzymes are different. I have recently established the NMR parameters for observing the paramagnetic relaxation effects of NOS with arginine and several inhibitors. Th paramagnetic effect of the Fe will be used to obtain distant constraints from the Fe to the proton of each inhibitor and therefore establish overall distance as well as conformational restraints within the active site. Our current work is directed toward obtaining data for all three isoforms with selected inhibitors. Computer Graphics Laboratory resources, including MidasPlus, is used to construct a model of each active site together with relative inhibitor affinities, toward the rational design of isofrom selective inhibitors.
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