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Electron-transfer-pathway state

From Bioblast


high-resolution terminology - matching measurements at high-resolution


Electron-transfer-pathway state

Description

Substrate control states are obtained in mitochondrial preparations (isolated mitochondria, permeabilized cells, permeabilized tissues, tissue homogenate) by depletion of endogenous substrates and addition of specific substrates to the mitochondrial respiration medium. Mitochondrial substrate control states have to be defined complementary to mitochondrial coupling control states. Coupling states (LEAK, OXPHOS, ETS) require electron transfer system competent substrate states, including oxygen supply. Two fundamental substrate control states can be distinguished (1) Substrate control states with electron entry through a single respiratory Complex (electron gating): Substrate control with electron entry separately through Complex I (pyruvate&malate or glutamate&malate) or Complex II (succinate and rotenone) restricts ETS capacity and artificially enhances flux control upstream of the Q-junction, providing diagnostic information on specific branches of the ETS. (2) Physiological substrate control states with convergent electron flow: Physiological combinations of Complex I&II substrates or even with additional convergent substrate supply through CGpDH and CETF support maximum ETS and OXPHOS capacities, due to the additive effect of multiple electron supply pathways converging at the Q-junction. Β» MiPNet article

Abbreviation: n.a.

Reference: Gnaiger 2009 Int J Biochem Cell Biol, Gnaiger 2014 MitoPathways


MitoPedia methods: Respirometry 


MitoPedia topics: "Respiratory state" is not in the list (Enzyme, Medium, Inhibitor, Substrate and metabolite, Uncoupler, Sample preparation, Permeabilization agent, EAGLE, MitoGlobal Organizations, MitoGlobal Centres, ...) of allowed values for the "MitoPedia topic" property. Respiratory state"Respiratory state" is not in the list (Enzyme, Medium, Inhibitor, Substrate and metabolite, Uncoupler, Sample preparation, Permeabilization agent, EAGLE, MitoGlobal Organizations, MitoGlobal Centres, ...) of allowed values for the "MitoPedia topic" property. 

Substrate control states

Publications in the MiPMap
Gnaiger E (2015) Substrate control states. Mitochondr Physiol Network 2015-03-10.


OROBOROS (2015) MiPNet

Abstract: Substrate control states are defined complementary to coupling control states in mitochondrial physiology.


β€’ O2k-Network Lab: AT Innsbruck Gnaiger E


Labels:



Preparation: Permeabilized cells, Permeabilized tissue, Homogenate, Isolated mitochondria, SMP 


Coupling state: LEAK, OXPHOS, ETS"ETS" is not in the list (LEAK, ROUTINE, OXPHOS, ET) of allowed values for the "Coupling states" property. 

HRR: Theory 


ETS competent substrate control states

Coupling states (LEAK, OXPHOS, ETS) require electron transfer system (ETS) competent substrate states, including sufficient oxygen supply. ETS competence of external substrates requires (i) transport of substrates across the inner mt-membrane or oxidation by dehydrogenases located on the outer face of the inner mt-membrane (e.g. glycerophosphate dehydrogenase complex, CGpDH), (ii) oxidation in the mt-matrix (TCA cycle dehydrogenases and other matrix dehydrogenases, e.g. mtGDH) or on the inner face of the inner mt-membrane (succinate dehydrogenase), (iii) oxidation of substrates without accumulation of inhibitory endproducts (e.g. oxaloacetate inhibiting succinate dehydrogenase; NADH and oxaloacetate inhibiting malate dehydrogenase), and (iv) electron transfer through the membrane-bound ETS (mETS). Endproducts must be either easily exported from the matrix across the inner mt-membrane (e.g. malate formed from succinate via fumarate), or metabolized in the TCA cycle (e.g. malate-derived oxaloacetate forming citrate in the presence of external pyruvate&malate).


Respiratory Complex-specific substrate control states

Respiratory Complex-specific substrate control states are substrate control states for selective entry of electron transfer through one particular respiratory Complex; for instance through CI (PM; GM; PGM with or without malonic acid: Gnaiger 2014 MitoPathways Chapter 3, CII (S(Rot)), CIV (AsTm: MiPNet06.06_ChemicalBackground).


Physiological substrate control states

Physiological substrate control states are substrate control states obtained in intact cells respiring on endogenous substrates or in media with physiological exogenous substrates, or designed for reconstitution of TCA cycle function in isolated mitochondria, permeabilized cells or permeabilized tissues. In all cases, electron flow converges at the Q-junction with multiple entry sites of electron transfer through CI&II, CI&II&FAO, CI&II&GpDH.


Pathway versus kinetic substrate control

Control by substrate type: pathway control states

A: Intact cells

  1. Endogenous substrate control: In intact cells, endogenous organic carbon substrates are mobilized in the cytosol as intermediary metabolites transported across the inner mitochondrial membrane and thus exerting control over mitochondrial respiration. If no organic carbon substrates are supplied in the incubation medium, then substrate control is entirely endogenous. Long-term incubation under such conditions leads to progressive states of depletion of endogenous substrates.
  2. Exogenous substrate control: Cells are grown in complex culture media with a variety of organic carbon substrates, and different exogenous substrate control states are achieved by variation of these substrates. Long-term incubation in closed systems without exchange of culture medium leads to progressive states of depletion of exogenous substrates. Incubation of cells in simple media allows for sequential titration of specific carbon substrates (e.g. glucose or fructose; lactate or glutamate) for the study of exogenous substrate control of respiration.

B: Mitochondrial preparations

Specific substrate-inhibitor combinations are selected to establish substrate states for (i) stimulating defined segments of the electron transfer system, or (ii) reconstitution of TCA cycle function.
  1. Substrate control states with electron gating: Specific substrate-inhibitor combinations are applied for selectively stimulating electron entry though CI, CII, or other branches converging at the Q-junction, particularly with fatty acids and alpha-glycerophosphate (CI-linked respiration, CII-linked respiration, etc.). The most commonly applied substrate states select for Complex I electron input (CI: pyruvate&malate, PM; glutamate&malate, GM), Complex II electron input (CII: succinate and rotenone, S(Rot)), or Complex IV electron input (CIV: ascorbate&TMPD(Ama)).
  2. Physiological substrate control states: Reconstitution of TCA cycle function requires CI&II-linked substrate combinations, such as PMS, GMS, or PGMS, applied simultaneously without inhibitor of any respiratory complexes.


Control by substrate concentration: kinetic control states

  1. Kinetic substrate or adenylate control: Kinetic studies with variation of a specific substrate (reduced substrate supplying electrons to the ETS; ADP, Pi; O2; cytochrome c) are analyzed by kinetic functions (e.g. hyperbolic), yielding kinetic parameters, such as Jmax, Km', c50 [Β΅M], or p50 [kPa].
  2. Kinetic inhibitor control: Kinetic studies with variation of a specific inhibitor yield apparent kinetic constants, such as the KI'.