The ratio between tetrahydrobiopterin and oxidized tetrahydrobiopterin analogues controls superoxide release from endothelial nitric oxide synthase: an EPR spin trapping study
Open accessAbstract
Augmentation of superoxide levels has been linked to impaired relaxation in hypertension, diabetes and hypercholesterolaemia. Purified endothelial nitric oxide synthase (eNOS) generates superoxide under limited availability of 5,6,7,8-tetrahydrobiopterin (BH4). Thus alterations in endothelial BH4 levels have been postulated to stimulate superoxide production from eNOS. This possibility was examined by determining the concentration-dependent effects of BH4, and its analogues, on superoxide formation by eNOS. Superoxide was quantified by EPR spin trapping, which is the only available technique to quantify superoxide from eNOS. Using 5-ethoxycarbonyl-5-methyl-pyrroline N-oxide, we show that only fully reduced BH4 diminished superoxide release from eNOS, with efficiency BH4>6-methyl-BH4>5-methyl-BH4. In contrast, partially oxidized BH4 analogues, 7,8-dihydrobiopterin (7,8-BH2) and sepiapterin had no effect. Neither l-arginine nor NG-nitro-l-arginine methyl ester (l-NAME) abolished superoxide formation. Together, BH4 and l-arginine stimulated ˙NO production at maximal rates of 148nmol/min per mg of protein. These results indicate that BH4 acts as a ‘redox switch’, decreasing superoxide release and enhancing ˙NO formation. This role was verified by adding 7,8-BH2 or sepiapterin to fully active eNOS. Both 7,8-BH2 and sepiapterin enhanced superoxide release while inhibiting ˙NO formation. Collectively, these results indicate that the ratio between oxidized and reduced BH4 metabolites tightly regulates superoxide formation from eNOS. The pathological significance of this scenario is discussed.
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Gene splicing appears to modulate NOX4 activation and play important roles in cardiovascular disease.
“In addition, gene splicing also appears to modulate NOX4 activation and play important roles in cardiovascular disease (971).”
From Sources of Vascular Nitric Oxide and Reactive Oxygen Species and Their Regulation (2019) · cites this paper as [971]
Splicing of NOX4 appears to influence cardiovascular disease, with a shorter protein (NOX4D) exerting beneficial signaling functions in the nucleus, whereas full-length NOX4 is overexpressed in ischemia with detrimental effects.
“For instance, splicing of NOX4 appears to influence cardiovascular disease, with a shorter protein (NOX4D) exerting beneficial signaling functions in the nucleus, whereas full-length NOX4 is overexpressed in ischemia with detrimental effects (971).”
From Sources of Vascular Nitric Oxide and Reactive Oxygen Species and Their Regulation (2019) · cites this paper as [971]
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- Authors
- VÁSQUEZ-VIVAR, J., MARTÁSEK, P., WHITSETT, J., JOSEPH, J., & KALYANARAMAN, B.
- Affiliations
- Biophysics Research Institute and Free Radical Research Center, Medical College of Wisconsin, Milwaukee, WI 53226, U.S.A.
- University of Texas Health Science Center, San Antonio, TX 78284-7760, U.S.A.
- Medical College of Wisconsin (US)
- Corresponding author
- WHITSETT, Jennifer
- JOSEPH, Joy
- KALYANARAMAN, Balaraman
- Venue
- Biochemical Journal
- Publisher
- Portland Press Ltd.
- Published
- 2002-03-08
- Volume
- 362
- Issue
- 3
- Pages
- 733-739
- ISSN
- 0264-6021 (ISSN-L)
- 1470-8728
- PMC
- PMC1222439
- OpenAlex
- W2106121696
- Cited by
- 287
- References
- 0
- Citations by year
- 2026: 2 · 2025: 3 · 2024: 7 · 2023: 5 · 2022: 8 · 2021: 9 · 2020: 5 · 2019: 4 · 2018: 5 · 2017: 10 · 2016: 11 · 2015: 9 · 2014: 21 · 2013: 18 · 2012: 23
- FWCI
- 10.59
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- Biochemical Journal (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institute of General Medical Sciences (R01GM052419, GM-52419)
- Topics
- Health Sciences › Medicine › Physiology › Nitric Oxide and Endothelin Effects
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Redox biology and oxidative stress
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Electron Spin Resonance Studies
- Keywords
- tetrahydrobiopterin, superoxide production, endothelial nitric oxide synthase, 7,8-dihydrobiopterin, sepiapterin, nitric oxide production, redox switch, EPR spin trapping, L-arginine, hypertension, endothelial dysfunction
- MeSH terms
- Arginine, Biopterins, Electron Spin Resonance Spectroscopy, Endothelium, Vascular, Kinetics, Oxidation-Reduction, Recombinant Proteins, Stereoisomerism, Structure-Activity Relationship, Superoxides, Nitric Oxide Synthase, NG-Nitroarginine Methyl Ester, Nitric Oxide Synthase Type III
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