Key Event Title
|Level of Biological Organization|
Key Event Components
Key Event Overview
AOPs Including This Key Event
|AOP Name||Role of event in AOP|
|Mitochondrial complex inhibition leading to liver injury||MolecularInitiatingEvent|
Key Event Description
The mitochondrial complex V (ATPase) is an enzyme complex located in the inner membrane of mitochondria. It is the fifth out of 5 complexes that together form the mitochondrial respiratory chain.
The ATPase catalysed the production of ATP from ADP and Pi using the proton gradient created by complex I,III and IV.
The initiation event is the reversibly or irreversibly interaction to any of the subunits in the mitochondrial complex V, leading to an perturbation of ATP production.
How It Is Measured or Detected
Complex inhibition assays specifically for complex V
The activity of complex IV is always tested in the bovine heart mitochondria provided within the kit or using freshly isolated mitochondria. The kit relies on the following detection of complex V activity:
ATP is converted by ATPase to ADP. The ADP is used by pyruvate kinase to convert Phosphoenolpyruvate to pyruvate. The pyruvate is converted to lactate by lactate dehydrogenase, while oxidising NADH to NAD+. NADH oxidation is measured at 340nm.
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
MitoCheck® Complex V Activity Assay Kit (Item No. 701000)
MitoTox™ Complex V OXPHOS Activity Assay Kit (ab109907)
Membrane potential dyes
Perry, 2011, mitochondrial membrane potential probes and proton gradient
Mitra, 2010, analysis of mitochondrial dynamics and functions using imaging approaches
Domain of Applicability
Evidence for Perturbation by Stressor
Overview for Molecular Initiating Event