State 2

high-resolution terminology - matching measurements at high-resolution

State 2

Description

Substrate limited state of residual oxygen consumption, after addition of ADP to isolated mitochondria suspended in mitochondrial respiration medium in the absence of reduced substrates (ROX_D). Residual endogenous substrates are oxidized during a transient stimulation of oxygen flux by ADP. The peak – supported by endogenous substrates – is, therefore, a pre-steady state phenomenon preceding State 2. Subsequently oxygen flux declines to a low level (or zero) at the steady State 2 (Chance and Williams 1955). ADP concentration (D) remains high during ROX_D.

Abbreviation: ROX_D

Reference: Chance 1955 JBC-III; MiPNet12.15; List of respiratory states

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. 

An alternative protocol

Sequential addition of (1) mitochondria, (2) ADP, and (3) reduced substrates is the basis of the original State 1-2-3 definitions of respiratory states (Chance and Williams 1955 part III, 1956), where State 2 is zero respiration or residual oxygen consumption in the absence of substrate. An alternative protocol is well established, as shown e.g. by the classical Fig. 5A (Chance and Williams 1955 part I): 600 µM ADP is added after a state described as ‘Aerobic mitochondria plus succinate’. That state was never defined as ‘State 2’ by Brit Chance. Later Estabrook (1967) made this protocol more popular, with addition of substrate before any ADP or ATP was added.

In this alternative protocol, a respiratory LEAK state is induced as the second respiratory state of isolated mitochondria, permeabilized tissues, or permeabilized cells, adding the mitochondrial preparation to respiration medium containing inorganic phosphate (State 1), then adding reduced substrate (no external adenylates). This second state (Estabrook 1967) is a non-phosphorylating LEAK state, L_N (N for no adenylates; Gnaiger 2009), when substrate-saturated respiration compensates for the proton leak (mainly) in the absence of ADP.

In contradiction to the original definition of State 2 (ROX), yet with reference to Chance and Williams (1956), 'State 2' has later been used for describing this functionally different state of LEAK respiration:

‘State 2: substrate added, respiration low due to lack of ADP. .. the controlled respiration prior to addition of ADP, which is strictly termed “state 2”, is functionally the same as state 4, and the latter term is usually used for both states’ (Nicholls & Ferguson 1992).

To overcome the termonological confusion persisting in the scientific literature, the respiratory coupling states of LEAK respiration, OXPHOS capacity and ETS capacity are distinguished from residual oxygen consumption (ROX; Gnaiger 2009).

References

Chance B, Williams GR (1955) Respiratory enzymes in oxidative phosphorylation. I. Kinetics of oxygen utilization. J. Biol. Chem. 217: 383-393.

Chance B, Williams GR (1955) Respiratory enzymes in oxidative phosphorylation. III. The steady state. J. Biol. Chem. 217: 409-427.

Chance B, Williams GR (1956) The respiratory chain and oxidative phosphorylation. Adv. Enzymol. 17: 65-134.

Estabrook R (1967) Mitochondrial respiratory control and the polarographic measurement of ADP:O ratios. Methods Enzymol. 10: 41-47.

Gnaiger E (2009) Capacity of oxidative phosphorylation in human skeletal muscle. New perspectives of mitochondrial physiology. Int. J. Biochem. Cell Biol. 41: 1837–1845. PMID: 19467914

Nicholls DG, Ferguson SJ (2002) Bioenergetics 3. Academic Press, London. 287 pp.