Linked Life Data

20184376

Source:http://linkedlifedata.com/resource/pubmed/id/20184376

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pubmed-article:20184376pubmed:abstractTextEndothelial nitric oxide synthase (eNOS) is an important regulator of vascular and cardiac function. Peroxynitrite (ONOO(-)) inactivates eNOS, but questions remain regarding the mechanisms of this process. It has been reported that inactivation is due to oxidation of the eNOS zinc-thiolate cluster, rather than the cofactor tetrahydrobiopterin (BH(4)); however, this remains highly controversial. Therefore, we investigated the mechanisms of ONOO(-)-induced eNOS dysfunction and their dose dependence. Exposure of human eNOS to ONOO(-) resulted in a dose-dependent loss of activity with a marked destabilization of the eNOS dimer. HPLC analysis indicated that both free and eNOS-bound BH(4) were oxidized during exposure to ONOO(-); however, full oxidation of protein-bound biopterin required higher ONOO(-) levels. Additionally, ONOO(-) triggered changes in the UV/visible spectrum and heme content of the enzyme. Preincubation of eNOS with BH(4) decreased dimer destabilization and heme alteration. Addition of BH(4) to the ONOO(-)-destabilized eNOS dimer only partially rescued enzyme function. In contrast to ONOO(-) treatment, incubation with the zinc chelator TPEN with removal of enzyme-bound zinc did not change the eNOS activity or stability of the SDS-resistant eNOS dimer, demonstrating that the dimer stabilization induced by BH(4) does not require zinc occupancy of the zinc-thiolate cluster. While ONOO(-) treatment was observed to induce loss of Zn binding, this cannot account for the loss of enzyme activity. Therefore, ONOO(-)-induced eNOS inactivation is primarily due to oxidation of BH(4) and irreversible destruction of the heme/heme center.lld:pubmed
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pubmed-article:20184376pubmed:authorpubmed-author:TsaiAh-LimALlld:pubmed
pubmed-article:20184376pubmed:authorpubmed-author:BerkaVladimir...lld:pubmed
pubmed-article:20184376pubmed:authorpubmed-author:ZweierJay LJLlld:pubmed
pubmed-article:20184376pubmed:authorpubmed-author:HemannCraigClld:pubmed
pubmed-article:20184376pubmed:authorpubmed-author:ChenWeiguoWlld:pubmed
pubmed-article:20184376pubmed:authorpubmed-author:ChenYeong-Ren...lld:pubmed
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