Malate

From Bioblast
Jump to: navigation, search
Bioblasts - Richard Altmann and MiPArt by Odra Noel
MitoPedia     Terms and abbreviations     Preprints and history     MiP and biochemistry     Concepts and methods     MitoPedia: SUIT     MitoPedia: O2k



MitoPedia

Malate

Description

Malic acid

Malic acid, C4H6O5, occurs under physiological conditions as the anion malate2-, M, with pKa1 = 3.40 and pKa2 = 5.20. L-Malate is formed from fumarate in the TCA cycle in the mitochondrial matrix, where it is the substrate of malate dehydrogenase oxidized to oxaloacetate. Malate is also formed in the cytosol. It cannot permeate through the lipid bilayer of membranes and hence requires a carrier (dicarboxylate carrier, tricarboxylate carrier and 2-oxoglutarate carrier). Malate alone cannot support respiration of mt-preparations from most tissues, since oxaloacetate accumulates in the absence of pyruvate or glutamate. Malate is a type N substrate (N) required for the FAO-pathway. In the presence of anaplerotic pathways (e.g., mitochondrial malic enzyme, mtME) FAO-pathway could be overestimated due to a contribution of NADH-linked respiration, F(N) (see SUIT-002).

Abbreviation: M

Reference: Gnaiger 2020 BEC MitoPathways

Application in HRR

M: Malate (L-Malic acid, C4H605); Sigma M 1000, 100 g, store at RT; M = 134.1 g·mol-1.


Preparation of 400 mM stock solution (dissolved in H2O):
  1. Weigh 268.2 mg of L-malic acid.
  2. Add 3 mL H2O.
  3. Neutralize with 5 M KOH (approx. 970 µL).
  4. Adjust final volume to 5 mL (in 5 mL volumetric glass flask).
  5. Divide into 0.5 mL portions.
  6. Store frozen at -20 °C.
»O2k manual titrations MiPNet09.12 O2k-Titrations
  • Titration volume (2-mL O2k-chamber): 10 µL using a 25 µL Hamilton syringe.
  • Titration volume (0.5-mL O2k-chamber): 2.5 µL using a 10 µL Hamilton syringe
  • Final concentration: 2 mM.


Preparation of 50 mM solution
  1. Dilute 0.625 mL of 400 mM stock solution with 4.375 ml of H2O (final volume 5 mL)
  2. Divide into 0.250 mL portions.
  3. Store frozen at -20 °C.
»O2k manual titrations MiPNet09.12 O2k-Titrations
  • Titration volume (2-mL O2k-chamber): 4 µL using a 10 µL Hamilton syringe
  • Titration volume (0.5-mL O2k-chamber): 1 µL using a 10 µL Hamilton syringe
  • Final concentration: 0.1 mM.


Bioblast wiki

Malate: 0.5 mM versus 2 mM in HRR

In mitochondrial preparations obtained from a diversity of tissues and organisms, malate at concentrations >0.5 mM exerted an inhibitory effect on succinate-pathway (S-pathway) and NADH and S (NS-)pathway, whereas 0.5 mM malate was apparently saturating for NADH (N-) linked respiration. However, this is not true in the presence of malonate inhibiting Complex II. Selection of an optimum malate concentration for SUIT protocols is a compromise. Current investigations point towards 2 mM malate as the optimum concentration. Sumbalova_2016a_Abstract_MitoFit_Science_Camp_2016
  • History
800 mM stock until 2013-11-20 (for 2 mM final concentration). During this year, test experiments with mitochondria from various tissues and species and with different mt-preparations (isolated mitochondria, permeabilized fibres, tissue homogenate) revealed an inhibitory effect of 2 mM malate on succinate-linked respiration at 10 mM succinate (S,Rot). The inhibitory effect is less at 0.5 mM malate (400 mM stock for 0.5 mM experimental concentration) and is reversed by 50 mM succinate.


MitoPedia topics: Substrate and metabolite