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Simultaneous Measurement of Superoxide/Hydrogen Peroxide and NADH Production by Flavin-containing Mitochondrial Dehydrogenases
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JoVE Journal Biochimica
Simultaneous Measurement of Superoxide/Hydrogen Peroxide and NADH Production by Flavin-containing Mitochondrial Dehydrogenases

Simultaneous Measurement of Superoxide/Hydrogen Peroxide and NADH Production by Flavin-containing Mitochondrial Dehydrogenases

DOI:
10.3791/56975-v

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08:57 min

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February 24, 2018

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, , ,
1Department of Biochemistry,Memorial University of Newfoundland

Capitoli

  • 00:00Titolo
  • 00:17Introduction
  • 01:50Setting Up the Assay
  • 05:44Results: Simultaneous tracking of changes in NADH and superoxide/hydrogen peroxide fluorescence
  • 07:32Conclusion

Summary

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Mitochondria contain several flavin-dependent enzymes that can produce reactive oxygen species (ROS). Monitoring ROS release from individual sites in mitochondria is challenging due to unwanted side reactions. We present an easy, inexpensive method for direct assessment of native rates for ROS release using purified flavoenzymes and microplate fluorometry.

Tags

Mitochondrial DehydrogenasesSuperoxide/hydrogen PeroxideNADH ProductionFlavin-containing EnzymesPyruvate DehydrogenaseAlpha-ketoglutarate DehydrogenaseMicroplate FluorometryReactive Oxygen SpeciesAntioxidantsInhibitorsSimultaneous Measurement

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