- 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.
- Substrate control states with electron entry through a single respiratory Complex (electron gating);
- Physiological substrate control states with convergent electron flow.
Abbreviation: n.a.
Reference: Gnaiger 2009 Int J Biochem Cell Biol, Gnaiger_2012_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.
Pathway versus kinetic substrate control
Author: Gnaiger E, OROBOROS INSTRUMENTS, Innsbruck, Austria. Last edit: 2012-12-02
Control by substrate type: Pathway control states
A: Intact cells
- 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.
- 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.
- 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.
- Substrate control states with electron gating: Specific substrate-inhibitor combinations are applied for selectively stimulating electron entry though Complex I, CII, or other branches converging at the Q-junction, particularly with fatty acids and alpha-glycerophosphate (CI respiration, CII 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+rotenone, S(Rot)), or Complex IV electron input (CIV: ascorbate+TMPD(Ama)).
- Physiological substrate control states: Reconstitution of TCA cycle function requires CI+II-linked substrate combinations, such as PMS, GMS, or PMGS, applied simultaneously without inhibitor of any respiratory complexes.
Control by substrate concentration: Kinetic control states
- 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 apparent kinetic constants, such as Jmax, Km', c50, or p50.
- Kinetic inhibitor control: Kinetic studies with variation of a specific inhibitor yield apparent kinetic constants, such as the KI'.
