Characterization of the magnitude and kinetics of xanthine oxidase-catalyzed nitrate reduction: evaluation of its role in nitrite and nitric oxide generation in anoxic tissues
Abstract
In addition to nitric oxide (NO) generation from specific NO synthases, NO is also formed during anoxia from nitrite reduction, and xanthine oxidase (XO) catalyzes this process. While in tissues and blood high nitrate levels are present, questions remain regarding whether nitrate is also a source of NO and if XO-mediated nitrate reduction can be an important source of NO in biological systems. To characterize the kinetics, magnitude, and mechanism of XO-mediated nitrate reduction under anaerobic conditions, EPR, chemiluminescence NO-analyzer, and NO-electrode studies were performed. Typical XO reducing substrates, xanthine, NADH, and 2,3-dihydroxybenz-aldehyde, triggered nitrate reduction to nitrite and NO. The rate of nitrite production followed Michaelis-Menten kinetics, while NO generation rates increased linearly following the accumulation of nitrite, suggesting stepwise-reduction of nitrate to nitrite then to NO. The molybdenum-binding XO inhibitor, oxypurinol, inhibited both nitrite and NO production, indicating that nitrate reduction occurs at the molybdenum site. At higher xanthine concentrations, partial inhibition was seen, suggesting formation of a substrate-bound reduced enzyme complex with xanthine blocking the molybdenum site. The pH dependence of nitrite and NO formation indicate that XO-mediated nitrate reduction occurs via an acid-catalyzed mechanism. With conditions occurring during ischemia, myocardial xanthine oxidoreductase and nitrate levels were determined to generate up to 20 microM nitrite within 10-20 min that can be further reduced to NO with rates comparable to those of maximally activated NOS. Thus, XOR catalyzed nitrate reduction to nitrite and NO occurs and can be an important source of NO production in ischemic tissues.
Integrity signals
Registry
Cites
Statistics
Claims the paper makes
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
Animals do not encode a classical nitrate reductase, although bovine xanthine oxidase can reduce nitrate into nitrite.
“Eukaryotic nitrate reductase (eukNR) has been isolated from plants and fungi; animals do not encode a classical nitrate reductase, although bovine xanthine oxidase (XO) can reduce nitrate into nitrite.”
From Nitrate and periplasmic nitrate reductases (2014) · cites this paper as [77]
The nitrite reductase activity of neuroglobin or cytochrome c is negligible when the heme is hexacoordinated.
“the nitrite reductase activity of neuroglobin [63, 64, 70, 76] or cytochrome c [47] is negligible when the heme is hexacoordinated.”
From Current perspectives and challenges in understanding the role of nitrite as an integral player in nitric oxide biology and therapy (2011) · cites this paper as [76]
XOR nitrate reductase activity is highly pH-dependent, with higher activity occurring at lower pH.
“It has been shown that XOR nitrate reductase activity is highly pH-dependent, with higher activity occurring at lower pH [12].”
From Skeletal muscle as an endogenous nitrate reservoir (2015) · cites this paper as [12]
Cites
Endothelium‐derived relaxing and contracting factors
Furchgott, R. F., & Vanhoutte, P. M. (1989) · The FASEB Journal · doi:10.1096/fasebj.3.9.2545495 · Cited by 2,007 · see citing works
Nitric oxide, a novel neuronal messenger
Bredt, D. S., & Snyder, S. H. (1992) · Neuron · doi:10.1016/0896-6273(92)90104-l · Cited by 1,852 · see citing works
Benjamin, N., O'Driscoll, F., Dougall, H., Duncan, C., Smith, L., Golden, M., & McKenzie, H. (1994) · Nature · doi:10.1038/368502a0 · Cited by 665 · see citing works
Millar, T. M., Stevens, C. R., Benjamin, N., Eisenthal, R., Harrison, R., & Blake, D. R. (1998) · FEBS Letters · doi:10.1016/s0014-5793(98)00430-x · Cited by 491 · see citing works
Reduction of nitrite to nitric oxide catalyzed by xanthine oxidoreductase
Godber, B. L. J., Doel, J. J., Sapkota, G. P., Blake, D. R., Stevens, C. R., Eisenthal, R., & Harrison, R. (2000) · Journal of Biological Chemistry · doi:10.1074/jbc.275.11.7757 · Cited by 411 · see citing works
Non-Enzymatic Nitric Oxide Synthesis in Biological Systems
Zweier, J. L., Samouilov, A., & Kuppusamy, P. (1999) · Biochimica et Biophysica Acta (BBA) - Bioenergetics · doi:10.1016/s0005-2728(99)00018-3 · Cited by 363 · see citing works
Self-citationZweier, J. L., Broderick, R., Kuppusamy, P., Thompson-Gorman, S. L., & Lutty, G. A. (1994) · Journal of Biological Chemistry · doi:10.1016/s0021-9258(19)51062-6 · Cited by 244 · see citing works
Self-citationSodium N‐Methyl‐D‐glucamine Dithiocarbamate and Cadmium Intoxication
Shinobu, L. A., Jones, S. G., & Jones, M. M. (1984) · Acta Pharmacologica et Toxicologica · doi:10.1111/j.1600-0773.1984.tb01916.x · Cited by 221 · see citing works
Giraldez, R. R., Panda, A., Xia, Y., Sanders, S. P., & Zweier, J. L. (1997) · Journal of Biological Chemistry · doi:10.1074/jbc.272.34.21420 · Cited by 180 · see citing works
Self-citationEvaluation of the Magnitude and Rate of Nitric Oxide Production from Nitrite in Biological Systems
Samouilov, A., Kuppusamy, P., & Zweier, J. L. (1998) · Archives of Biochemistry and Biophysics · doi:10.1006/abbi.1998.0785 · Cited by 171 · see citing works
Self-citationEvaluation of the role of xanthine oxidase in myocardial reperfusion injury.
Thompson-Gorman, S. L., & Zweier, J. L. (1990) · Journal of Biological Chemistry · doi:10.1016/s0021-9258(19)39200-2 · Cited by 168 · see citing works
Self-citationEffect of ischemia reperfusion or hypoxia reoxygenation on lung vascular permeability and resistance
Allison, R. C., Kyle, J. C., Adkins, W. K., Prasad, V. R., McCord, J., & Taylor, A. E. (1990) · Journal of Applied Physiology · doi:10.1152/jappl.1990.69.2.597 · Cited by 94 · see citing works
Dixon, M., & Thurlow, S. (1924) · Biochemical Journal · doi:10.1042/bj0180971 · Cited by 84 · see citing works
The inhibition of xanthine oxidase by 8-bromoxanthine.
Hille, R., & Stewart, R. C. (1984) · Journal of Biological Chemistry · doi:10.1016/s0021-9258(17)43447-8 · Cited by 52 · see citing works
Substrate inhibition of xanthine oxidase and its influence on superoxide radical production
Rubbo, H., Radí, R., & Prodanov, E. (1991) · Biochimica et Biophysica Acta (BBA) - General Subjects · doi:10.1016/0304-4165(91)90089-y · Cited by 33 · see citing works
Flagged first, then by citations
Cited by
Sorted by citations
Nitrite reduction to nitric oxide by deoxyhemoglobin vasodilates the human circulation
Cosby, K., Partovi, K. S., Crawford, J. H., Patel, R. P., Reiter, C. D., Martyr, S., Yang, B. K., Waclawiw, M. A., Zalos, G., Xu, X., et al. (2003) · Nature Medicine · doi:10.1038/nm954 · Cited by 1,807 · see citing works
The mononuclear molybdenum enzymes
Hille, R., Hall, J., & Basu, P. (2014) · Chemical Reviews · doi:10.1021/cr400443z · Cited by 1,738 · see citing works
Reactive oxygen species in cardiovascular disease
Sugamura, K., & Keaney (2011) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2011.05.004 · Cited by 769 · see citing works
Cytoprotective effects of nitrite during in vivo ischemia-reperfusion of the heart and liver
Duranski, M. R., Greer, J. J. M., Dejam, A., Jaganmohan, S., Hogg, N., Langston, W., Patel, R. P., Yet, S.-F., Wang, X., Kevil, C. G., et al. (2005) · Journal of Clinical Investigation · doi:10.1172/jci22493 · Cited by 674 · see citing works
Plasma nitrite reflects constitutive nitric oxide synthase activity in mammals
Kleinbongard, P., Dejam, A., Lauer, T., Rassaf, T., Schindler, A., Picker, O., Scheeren, T., Gödecke, A., Schrader, J., Schulz, R., et al. (2003) · Free Radical Biology and Medicine · doi:10.1016/s0891-5849(03)00406-4 · Cited by 586 · see citing works
Govoni, M., Jansson, E. Å., Weitzberg, E., & Lundberg, J. O. (2008) · Nitric Oxide · doi:10.1016/j.niox.2008.08.003 · Cited by 585 · see citing works
Nitrite as Regulator of Hypoxic Signaling in Mammalian Physiology
van Faassen, E. E., Bahrami, S., Feelisch, M., Hogg, N., Kelm, M., Kim‐Shapiro, D. B., Kozlov, A. V., Li, H., Lundberg, J. O., Mason, R., et al. (2009) · Medicinal Research Reviews · doi:10.1002/med.20151 · Cited by 435 · see citing works
Self-citationNitrate and periplasmic nitrate reductases
Sparacino-Watkins, C., Stolz, J. F., & Basu, P. (2014) · Chem. Soc. Rev. · doi:10.1039/c3cs60249d · Cited by 426 · see citing works
Hendgen-Cotta, U. B., Merx, M. W., Shiva, S., Schmitz, J., Becher, S., Klare, J. P., Steinhoff, H.-J., Goedecke, A., Schrader, J., Gladwin, M. T., et al. (2008) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.0801336105 · Cited by 418 · see citing works
Maia, L. B., & Moura, J. J. G. (2014) · Chemical Reviews · doi:10.1021/cr400518y · Cited by 401 · see citing works
A mammalian functional nitrate reductase that regulates nitrite and nitric oxide homeostasis
Jansson, E. Å., Huang, L., Malkey, R., Govoni, M., Nihlén, C., Olsson, A., Stensdotter, M., Petersson, J., Holm, L., Weitzberg, E., et al. (2008) · Nature Chemical Biology · doi:10.1038/nchembio.92 · Cited by 356 · see citing works
NO Generation From Nitrite and Its Role in Vascular Control
Lundberg, J. O., & Weitzberg, E. (2005) · Arteriosclerosis, Thrombosis, and Vascular Biology · doi:10.1161/01.atv.0000161048.72004.c2 · Cited by 341 · see citing works
Ferguson, S. K., Hirai, D. M., Copp, S. W., Holdsworth, C. T., Allen, J. D., Jones, A. M., Musch, T. I., & Poole, D. C. (2013) · The Journal of Physiology · doi:10.1113/jphysiol.2012.243121 · Cited by 333 · see citing works
Kapil, V., Khambata, R. S., Jones, D. A., Rathod, K., Primus, C., Massimo, G., Fukuto, J. M., & Ahluwalia, A. (2020) · Pharmacological Reviews · doi:10.1124/pr.120.019240 · Cited by 248 · see citing works
Fructose Intake, Serum Uric Acid, and Cardiometabolic Disorders: A Critical Review
Caliceti, C., Calabria, D., Roda, A., & Cicero, A. (2017) · Nutrients · doi:10.3390/nu9040395 · Cited by 236 · see citing works
Lang, J. D., Teng, X., Chumley, P., Crawford, J. H., Isbell, T. S., Chacko, B. K., Liu, Y., Jhala, N., Crowe, D. R., Smith, A. B., et al. (2007) · Journal of Clinical Investigation · doi:10.1172/jci31892 · Cited by 224 · see citing works
Nitrite: A physiological store of nitric oxide and modulator of mitochondrial function
Shiva, S. (2013) · Redox Biology · doi:10.1016/j.redox.2012.11.005 · Cited by 207 · see citing works
“While the exact reaction scheme underlying XOR-mediated NO2 reduction is unclear, it is known that this reaction occurs at the molybdenum cofactor of XOR and aldehyde oxidase [11,12].”
BackgroundMechanisms of nitrite bioactivation
Kim-Shapiro, D. B., & Gladwin, M. T. (2014) · Nitric Oxide · doi:10.1016/j.niox.2013.11.002 · Cited by 157 · see citing works
Jones, A. M., Vanhatalo, A., Seals, D. R., Rossman, M. J., Piknova, B., & Jonvik, K. L. (2021) · Medicine & Science in Sports & Exercise · doi:10.1249/mss.0000000000002470 · Cited by 156 · see citing works
Characterization of the effects of oxygen on xanthine oxidase-mediated nitric oxide formation
Li, H., Samouilov, A., Liu, X., & Zweier, J. L. (2004) · Journal of Biological Chemistry · doi:10.1074/jbc.m314336200 · Cited by 146 · see citing works
Self-citationLi, H., Hemann, C., Abdelghany, T. M., El-Mahdy, M. A., & Zweier, J. L. (2012) · Journal of Biological Chemistry · doi:10.1074/jbc.m112.342378 · Cited by 132 · see citing works
Self-citationMechanisms of nitrite reduction to nitric oxide in the heart and vessel wall
Zweier, J. L., Li, H., Samouilov, A., & Liu, X. (2010) · Nitric Oxide · doi:10.1016/j.niox.2009.12.004 · Cited by 132 · see citing works
Self-citationLi, H., Kundu, T. K., & Zweier, J. L. (2009) · Journal of Biological Chemistry · doi:10.1074/jbc.m109.019125 · Cited by 126 · see citing works
Skeletal muscle as an endogenous nitrate reservoir
Piknova, B., Park, J. W., Swanson, K. M., Dey, S., Noguchi, C. T., & Schechter, A. N. (2015) · Nitric Oxide · doi:10.1016/j.niox.2015.02.145 · Cited by 108 · see citing works
Al-Shehri, S. S., Duley, J. A., & Bansal, N. (2020) · Redox Biology · doi:10.1016/j.redox.2020.101524 · Cited by 82 · see citing works
Effects of nitrite on modulating ROS generation following ischemia and reperfusion
Raat, N. J. H., Shiva, S., & Gladwin, M. T. (2009) · Advanced Drug Delivery Reviews · doi:10.1016/j.addr.2009.02.002 · Cited by 79 · see citing works
Nitrite reduction by xanthine oxidase family enzymes: a new class of nitrite reductases
Maia, L. B., & Moura, J. J. G. (2011) · JBIC Journal of Biological Inorganic Chemistry · doi:10.1007/s00775-010-0741-z · Cited by 75 · see citing works
Nitrate as a source of nitrite and nitric oxide during exercise hyperemia in rat skeletal muscle
Piknova, B., Park, J. W., Kwan (Jeff) Lam, K., & Schechter, A. N. (2016) · Nitric Oxide · doi:10.1016/j.niox.2016.03.005 · Cited by 75 · see citing works
Chlorella sp.-Ameliorated Undesirable Microenvironment Promotes Diabetic Wound Healing
Wu, H., Yang, P., Li, A., Jin, X., Zhang, Z., & Lv, H. (2023) · Acta Pharmaceutica Sinica B · doi:10.1016/j.apsb.2022.06.012 · Cited by 65 · see citing works
Vitturi, D. A., & Patel, R. P. (2011) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2011.05.037 · Cited by 59 · see citing works
On the mechanism by which dietary nitrate improves human skeletal muscle function
Affourtit, C., Bailey, S. J., Jones, A. M., Smallwood, M. J., & Winyard, P. G. (2015) · Frontiers in Physiology · doi:10.3389/fphys.2015.00211 · Cited by 58 · see citing works
Mikula, I., Durocher, S., Martasek, P., Mutus, B., & Slama-Schwok, A. (2009) · Biochemical Journal · doi:10.1042/bj20080987 · Cited by 51 · see citing works
Hendgen-Cotta, U. B., Kelm, M., & Rassaf, T. (2010) · Nitric Oxide · doi:10.1016/j.niox.2009.10.003 · Cited by 46 · see citing works
Magnetic resonance study of the transmembrane nitrite diffusion
Samouilov, A., Woldman, Y. Y., Zweier, J. L., & Khramtsov, V. V. (2007) · Nitric Oxide · doi:10.1016/j.niox.2006.12.006 · Cited by 46 · see citing works
Xanthine Oxidase Inhibitors in Heart Failure: Where Do We Go From Here?
Tamariz, L., & Hare, J. M. (2015) · Circulation · doi:10.1161/circulationaha.115.016379 · Cited by 39 · see citing works
Rafikov, R., Rafikova, O., Aggarwal, S., Gross, C., Sun, X., Desai, J., Fulton, D., & Black, S. M. (2013) · Journal of Biological Chemistry · doi:10.1074/jbc.m112.423269 · Cited by 36 · see citing works
Liu, M., Zollbrecht, C., Peleli, M., Lundberg, J. O., Weitzberg, E., & Carlström, M. (2015) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2015.03.025 · Cited by 30 · see citing works
Quantitative aspects of nitric oxide production in the heart
Ghasemi, A., & Jeddi, S. (2022) · Molecular Biology Reports · doi:10.1007/s11033-022-07889-x · Cited by 16 · see citing works
Corti, P., Tejero, J., & Gladwin, M. T. (2013) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2013.09.020 · Cited by 13 · see citing works
- Authors
- Li, H., Samouilov, A., Liu, X., & Zweier, J. L.
- Affiliations
- Johns Hopkins University (US)
- Corresponding author
- Zweier, Jay L.
- Venue
- Biochemistry
- Publisher
- American Chemical Society (ACS)
- Published
- 2003-01-04
- Volume
- 42
- Issue
- 4
- Pages
- 1150-1159
- ISSN
- 0006-2960 (ISSN-L)
- 1520-4995
- OpenAlex
- W2027439180
- Cited by
- 180
- References
- 15
- Citations by year
- 2026: 1 · 2025: 2 · 2024: 9 · 2023: 2 · 2022: 5 · 2021: 11 · 2020: 7 · 2019: 5 · 2018: 5 · 2017: 9 · 2016: 8 · 2015: 12 · 2014: 6 · 2013: 11 · 2012: 8
- FWCI
- 6.13
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Biochemistry (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Health Sciences › Medicine › Physiology › Nitric Oxide and Endothelin Effects
- Health Sciences › Medicine › Nephrology › Gout, Hyperuricemia, Uric Acid
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Redox biology and oxidative stress
- Keywords
- xanthine oxidase, nitrate reduction, nitric oxide production, nitrite production, anoxia, ischemia, Michaelis-Menten kinetics, xanthine, NADH, 2,3-dihydroxybenzaldehyde
- MeSH terms
- Animals, Hypoxia, Catalysis, Electrochemistry, Electron Spin Resonance Spectroscopy, Kinetics, Luminescent Measurements, Male, Myocardium, Nitrates, Nitric Oxide, Nitrites, Nitrogen, Nitrogen Isotopes, Oxidation-Reduction, Xanthine Oxidase, Rats, Sprague-Dawley, Spin Trapping, Xanthine, Rats
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
Are you an author, or do you see a problem with this page? Request a review
