Catalase

From Bioblast


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Catalase

Description

Catalase catalyzes the dismutation of hydrogen peroxide to water and oxygen. Perhaps all cells have catalase, but mitochondria of most cells lack catalase. Cardiac mitochondria are exceptional in having mt-catalase activity (rat heart mitochondria: Radi et al 1991; mouse heart mitochondria: Rindler et al 2013). Hydroxylamine is an inhibitor of catalase, which is also inhibited by cyanide and azide.

Mitochondrial respiration medium MiR05 was developed considering the intracellular conditions of mitochondria in living cells. In mitochondrial preparations, enzymes and substrates present in the cytosol (such as catalase) are diluted when the plasma membrane is removed. Therefore, the addition of catalase is recommended when working with mitochondrial preparations, to consume any H2O2 generated during the assay.

Abbreviation: Ctl

Reference: Radi 1991 J Biol Chem, Rindler 2013 J Biol Chem, MiPNet09.12

Communicated by Gnaiger E (2015-03-05) - last update with Timon-Gomez A (2023-07-13).

Application in HRR

Ctl: Catalase (from bovine liver, lyophilized powder, H2O2:H2O2 oxidoreductase; 2000-5000 U/mg protein), Sigma-Aldrich: C9322, store at -20 Β°C, CAS: 9001-05-2,
Hazard statements: H334; respiratory sensitization
Catalase is included in the mitochondrial respiration medium MiR06 (MiR06 = MiR05 plus catalase), so as to allow for re-oxygenation of the respiration medium (without lifting the stopper and risking the introduction of air bubbles) by addition of a small volume of a 200 mM stock solution of H2O2.
Unit definition: * Units of enzymatic activity (U) [Β΅mol/min]; assay used by Sigma Aldrich: ' One unit will decompose 1.0 ΞΌmole of H2O2 per min at pH 7.0 at 25 Β°C, while the H2O2 concentration falls from 10.3 to 9.2 mM, measured by the rate of decrease of A240. '
Use 'units/mg solid' for your calculations.
Example: 'Catalase lypophilized powder; 2800 U/mg solid and 3500 units/mg protein'.


Preparation of stock solution: 112 000 U/mL (dissolved in MiR05) for 2-mL O2k-chamber:
  1. Result: 40 mg catalase powder (2800 U/mg) are dissolved in 1 mL MiR05 to obtain a catalase stock solution with 112 000 U/mL.
  2. Divide into 0.2 mL portions.
  3. Store frozen at -20 Β°C.
Β»O2k manual titrations: MiPNet09.12
  • Titration volume (2-mL O2k-chamber): 5 Β΅L using a 25 Β΅L Hamilton syringe.
  • Final concentration: 280 U/mL.


Preparation of stock solution: 28 000 U/mL (dissolved in MiR05) for 0.5-mL O2k-chamber:
  1. 10 mg catalase powder (2800 U/mg) are dissolved in 1 mL MiR05 to obtain a catalase stock solution with 28 000 U/mL.
  2. Divide into 0.2 mL portions.
  3. Store frozen at -20 Β°C.
Β»O2k manual titrations: MiPNet09.12
  • Titration volume (0.5-mL O2k-chamber): 5 Β΅L using a 10 Β΅L Hamilton syringe.
  • Final concentration: 280 U/mL.


MitoPedia topics: Enzyme 

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