Cytochrome c

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Bioblasts - Richard Altmann and MiPArt by Odra Noel
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Cytochrome c

Description

Cytochrome c is a component of the Electron transfer-pathway (Electron transfer pathway) in mitochondria. It is a small heme protein loosely associated with the outer side of the inner mitochondrial membrane. The heme group of cytochrome c transfers electrons from Complex III to Complex IV. The release of cytochrome c into the cytoplasm is associated with apoptosis. Cytochrome c is applied in HRR to test the integrity of the mitochondrial outer membrane (cytochrome c control efficiency).

Abbreviation: c

Reference: MiPNet09.12; Gnaiger 2002 Biochem Soc Trans

Application in HRR: storage and stock solution of c

c: Cytochrome c (from equine heart; small hemeprotein), Sigma-Aldrich: C7752, store at -20 °C, CAS: 9007-43-6, M = 12.270 g·mol-1
The cytochrome c recommended (C7752, cytochrome c from equine heart) is out of stock for the next months (estimated delivery on November 25, 2021). The use of other cytochrome c sources is discussed in the MiPNet discussion forum.


Preparation of 4 mM cytochrome c solution (dissolved in H2O) for 2-mL O2k-chamber:
  1. Weigh 50 mg cytochrome c into a small glass beaker. Difficult to weigh, since the powder is electrostatically charged.
  2. Add 1 mL H2O; c dissolves easily.
  3. Divide into 0.2 mL portions.
  4. Store at -20 °C.
» O2k manual titrations: MiPNet09.12 O2k-Titrations
  • Titration volume (2-mL O2k-chamber): 5 µL using a 25 µL Hamilton syringe.
  • Final concentration: 10 µM.


Preparation of 5 mM cytochrome c solution (dissolved in H2O) for 0.5-mL O2k-chamber:
  1. Weigh 62,5 mg cytochrome c into a small glass beaker. Difficult to weigh, since the powder is electrostatically charged.
  2. Add 1 mL H2O; c dissolves easily.
  3. Divide into 0.2 mL portions.
  4. Store at -20 °C.
» O2k manual titrations: MiPNet09.12 O2k-Titrations
  • Titration volume (0.5-mL O2k-chamber): 1 µL using a 10 µL Hamilton syringe.
  • Final concentration: 10 µM.


DatLab oxygen flux: performance and data analysis

Quality of the results are strongly affected by the performance and data analysis. By adding cytochrome c in respirometric experiments the outer mitochondrial membrane integrity can be evaluated (cytochrome c control efficiency). The following DatLab traces illustrate examples of cytochrome c addition:
See also: Steady state, Figures 1-3.



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SUITbrowser question: mt outer membrane integrity

The cytochrome c test is available at several SUIT protocols. The SUITbrowser shows which protocols contain this test, alongside answer other research questions.




MitoPedia topics: Substrate and metabolite