Despite the considerable interest in superoxide as a potential cause of pathology, the mechanisms of its deleterious production by mitochondria remain poorly understood. Previous studies in purified mitochondria have found that the highest rates of superoxide production are observed with succinate-driven reverse-electron transfer through complex I, although the physiological importance of this pathway is disputed because it necessitates high concentrations of succinate and is thought not to occur when NAD is in the reduced state. However, very few studies have examined the rates of superoxide production with mitochondria respiring on both NADH-linked (e.g. glutamate) and complex II-linked substrates. In the present study, we find that the rates of superoxide production (measured indirectly as H2O2) with glutamate+succinate (∼1100 pmol of H2O2·min−1·mg−1) were unexpectedly much higher than with succinate (∼400 pmol of H2O2·min−1·mg−1) or glutamate (∼80 pmol of H2O2·min−1·mg−1) alone. Superoxide production with glutamate+succinate remained high even at low substrate concentrations (<1 mM), was decreased by rotenone and was completely eliminated by FCCP (carbonyl cyanide p-trifluoromethoxyphenylhydrazone), indicating that it must in large part originate from reverse-electron transfer through complex I. Similar results were obtained when glutamate was replaced with pyruvate, α-ketoglutarate or palmitoyl carnitine. In contrast, superoxide production was consistently lowered by the addition of malate (malate+succinate ∼30 pmol of H2O2·min−1·mg−1). We propose that the inhibitory action of malate on superoxide production can be explained by oxaloacetate inhibition of complex II. In summary, the present results indicate that reverse-electron transfer-mediated superoxide production can occur under physiologically realistic substrate conditions and suggest that oxaloacetate inhibition of complex II may be an adaptive mechanism to minimize this.
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Research Article|
December 21 2007
High rates of superoxide production in skeletal-muscle mitochondria respiring on both complex I- and complex II-linked substrates
Florian L. Muller;
Florian L. Muller
1
*Barshop Institute for Longevity and Aging Studies, University of Texas Health Science Center at San Antonio, Texas Research Park Campus, 15355 Lambda Drive, San Antonio, TX 78229-3900, U.S.A.
1To whom correspondence should be addressed (email aettius@aol.com).
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Yuhong Liu;
Yuhong Liu
†Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, U.S.A.
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Muhammad A. Abdul-Ghani;
Muhammad A. Abdul-Ghani
‡South Texas Veterans Health Care System, San Antonio, TX 78284-7762, U.S.A.
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Michael S. Lustgarten;
Michael S. Lustgarten
§Department of Physiology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, U.S.A.
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Arunabh Bhattacharya;
Arunabh Bhattacharya
*Barshop Institute for Longevity and Aging Studies, University of Texas Health Science Center at San Antonio, Texas Research Park Campus, 15355 Lambda Drive, San Antonio, TX 78229-3900, U.S.A.
†Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, U.S.A.
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Youngmok C. Jang;
Youngmok C. Jang
†Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, U.S.A.
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Holly Van Remmen
Holly Van Remmen
*Barshop Institute for Longevity and Aging Studies, University of Texas Health Science Center at San Antonio, Texas Research Park Campus, 15355 Lambda Drive, San Antonio, TX 78229-3900, U.S.A.
†Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, U.S.A.
‡South Texas Veterans Health Care System, San Antonio, TX 78284-7762, U.S.A.
§Department of Physiology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, U.S.A.
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Publisher: Portland Press Ltd
Received:
August 23 2007
Revision Received:
October 04 2007
Accepted:
October 05 2007
Accepted Manuscript online:
October 05 2007
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem J (2008) 409 (2): 491–499.
Article history
Received:
August 23 2007
Revision Received:
October 04 2007
Accepted:
October 05 2007
Accepted Manuscript online:
October 05 2007
Citation
Florian L. Muller, Yuhong Liu, Muhammad A. Abdul-Ghani, Michael S. Lustgarten, Arunabh Bhattacharya, Youngmok C. Jang, Holly Van Remmen; High rates of superoxide production in skeletal-muscle mitochondria respiring on both complex I- and complex II-linked substrates. Biochem J 15 January 2008; 409 (2): 491–499. doi: https://doi.org/10.1042/BJ20071162
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