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Key Event Title
Mitochondrial Complex IV inhibition
Key Event Components
Key Event Overview
AOPs Including This Key Event
|AOP Name||Role of event in AOP||Point of Contact||Author Status||OECD Status|
|Mitochondrial complex inhibition leading to liver injury||MolecularInitiatingEvent||Wanda van der Stel (send email)||Under development: Not open for comment. Do not cite|
Key Event Description
The mitochondrial complex IV (Cytochrome c oxidase) is an enzyme complex located in the inner membrane of mitochondria. It is the forth out of 5 complexes that together form the mitochondrial respiratory chain. It consists out of multiple subunits of cytochrome c oxidase, including cytochrome a and cytochrome a3.
The combination of these subunits catalyze following process:
The cytochrome c oxidase is involved in the process of oxidizing reduced cytochrome c to its oxidized form. A process that results in a free electron.
4 oxidation rounds results in enough electron to reduce 1 molecule oxygen to 2 molecules water.
The electron transfer in this process mediates the translocation of protons from the mitochondrial matrix through the inner membrane to the intermembrane space. The created proton gradient will be used to catalyze the reaction in which ADP is converted into ATP.
The initiation event is the reversibly or irreversibly interaction to any of the subunits in the mitochondrial complex IV, leading to an perturbation of the electron flow and an absence of proton transport via this complex.
How It Is Measured or Detected
Complex inhibition assays specifically for complex IV The activity of complex IV is always tested in the bovine heart mitochondria provided within the kit. The kit relies on the following detection of complex IV activity: The reduction of cytochrome c, which leads to a reduction in absorbance at 550nm can be measured.
Mitochondrial Membrane potential using fluorescent dyes. Positive charged molecules will accumulated in the mitochondria in an inverse proportion as the membrane potential. More polarised mitochondria will accumulate more dye (Rhodamine123, TMRE, TMRM) - leading to a higher fluorescent signal - and the absence of membrane potential leads to an absence of fluorescent signal. An exception is the dye JC1, because this dye has green fluorescence when present in low concentrations (depolarisation) and red fluorescence when accumulated (hyperpolarized
Complex inhibition assays
Cayman MitoCheck MitoCheck® Complex IV Activity Assay Kit (Item No. 700990)
Abcam MitoTox™ Complex IV OXPHOS Activity Assay Kit (ab109906) Complex IV Human Enzyme Activity Microplate Assay Kit (ab109909)
BioVision Cytochrome Oxidase Activity Colorimetric Assay Kit (K287)
Membrane potential dyes
Perry, 2011, mitochondrial membrane potential probes and proton gradient
Mitra, 2010, analysis of mitochondrial dynamics and functions using imaging approaches