S-glutathionylation uncouples eNOS and regulates its cellular and vascular function

Chun An Chen, Tse Yao Wang, Saradhadevi Varadharaj, Levy A. Reyes, Craig Hemann, M. A Hassan Talukder, Yeong Renn Chen, Lawrence J. Druhan, Jay L. Zweier

Research output: Contribution to journalArticlepeer-review

375 Scopus citations


Endothelial nitric oxide synthase (eNOS) is critical in the regulation of vascular function, and can generate both nitric oxide (NO) and superoxide (O2-2), which are key mediators of cellular signalling. In the presence of Ca2+/calmodulin, eNOS produces NO, endothelialderived relaxing factor, from L-arginine (L-Arg) by means of electron transfer from NADPH through a flavin containing reductase domain to oxygen bound at the haem of an oxygenase domain, which also contains binding sites for tetrahydrobiopterin (BH4) and L-Arg1-3. In the absence of BH4, NO synthesis is abrogated and instead O2-2 is generated4-7. While NOS dysfunction occurs in diseases with redox stress,BH4 repletion only partly restoresNOS activity and NOS-dependent vasodilation7. This suggests that there is an as yet unidentified redox-regulated mechanism controlling NOS function. Protein thiols can undergo S-glutathionylation, a reversible protein modification involved in cellular signalling and adaptation8,9. Under oxidative stress, S-glutathionylation occurs through thiol-disulphide exchange with oxidized glutathione or reaction of oxidant-induced protein thiyl radicals with reduced glutathione10,11. Cysteine residues are critical for the maintenance of eNOS function12,13; we therefore speculated that oxidative stress could alter eNOS activity through S-glutathionylation. Here we show that S-glutathionylation of eNOS reversibly decreases NOS activity with an increase in O2-2 generation primarily from the reductase, in which two highly conserved cysteine residues are identified as sites of S-glutathionylation and found to be critical for redox-regulation of eNOS function. We show that eNOS Sglutathionylation in endothelial cells, with loss of NO and gain of O2-2generation, is associated withimpaired endothelium-dependent vasodilation. In hypertensive vessels, eNOS S-glutathionylation is increased with impaired endothelium-dependent vasodilation that is restored by thiol-specific reducing agents, which reverse this S-glutathionylation. Thus, S-glutathionylation of eNOS is a pivotal switch providing redox regulation of cellular signalling, endothelial function and vascular tone.

Original languageEnglish (US)
Pages (from-to)1115-1120
Number of pages6
Issue number7327
StatePublished - Dec 23 2010
Externally publishedYes

ASJC Scopus subject areas

  • General


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