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Electron spin resonance spin-trapping detection of superoxide generated by neuronal nitric oxide synthase. Methods Enzymol 1999;301:169-77

Date

01/27/1999

Pubmed ID

9919565

DOI

10.1016/s0076-6879(99)01080-0

Scopus ID

2-s2.0-0031795460   36 Citations

Abstract

NOS is a ubiquitous enzyme that has an oxygenase and reductase activity. NOS reduces electron acceptors, at the reductase domain, by a one-electron mechanism that is not inhibited by SOD. One example of this activity is the direct reduction of ferricytochrome c by nNOS. Redox cycling electron acceptors (EA in Scheme 1), such as lucigenin and NBT, are reduced by NOS to generate an intermediate radical (EAred). This radical can then be reoxidized to the parent compound by oxygen, and in the process generate superoxide. Consequently, both NBT and lucigenin will enhance NADPH-dependent superoxide generation in the presence of flavoprotein reductases such as NOS. The artificial generation of superoxide from lucigenin and NBT is a major pitfall in the use of these compounds as superoxide probes. We conclude that the use of ESR spin-trapping techniques, although not free of problems, is a viable technique for the detection and quantification of superoxide in systems containing nNOS.

Author List

Vásquez-Vivar J, Martásek P, Hogg N, Karoui H, Masters BS, Pritchard KA Jr, Kalyanaraman B

Authors

Neil Hogg PhD Associate Dean, Professor in the Biophysics department at Medical College of Wisconsin
Balaraman Kalyanaraman PhD Chair, Professor in the Biophysics department at Medical College of Wisconsin
Kirkwood A. Pritchard PhD Professor in the Surgery department at Medical College of Wisconsin
Jeannette M. Vasquez-Vivar PhD Professor in the Biophysics department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Acridines
Animals
Electron Spin Resonance Spectroscopy
Free Radicals
Humans
Nitric Oxide Synthase
Spin Trapping
Superoxides
jenkins-FCD Prod-482 91ad8a360b6da540234915ea01ff80e38bfdb40a