State 4 is the respiratory state obtained in isolated mitochondria after State 3, when added ADP is phosphorylated maximally to ATP driven by electron transfer from defined respiratory substrates to O2 (Chance and Williams, 1955). State 4 represents LEAK respiration, LT (L for LEAK respiration; T for ATP), or an overestimation of LEAK respiration if ATPase activity prevents final accumulation of ATP and maintains a continuous stimulation of respiration by recycled ADP. This can be tested by inhibition of ATP synthase by oligomycin; LOmy). In the LEAK state (state of non-phosphorylating resting respiration; static head), oxygen flux is decreased to a minimum (corrected for ROX), and the mt-membrane potential is increased to a maximum for a specific substrate or substrate combination.
MitoPedia topics: EAGLE
Communicated by Gnaiger E 2010-08-15, edited 2016-03-20.
- State 4 is fundamentally different from State 2 (Chance and Williams, 1955), but unfortunately the latter has been misunderstood and re-defined to be comparable with State 4: ‘.. 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 and Ferguson 1992). The comparable states are not State 4 and State 2, but the LEAK state in the presence of ATP (State 4; LT) and in the absence of ATP and other adenylates (LN; N for no adenylates).
- State 4 yields identical respiratory flux as State 4o (Lo) only, if inhibition of ATP synthase by oligomycin (4o) is without effect compared to LT or LN.
- In living cells neither State 3 nor State 4 can be induced. (1) Exogenous substrates (if present) combine with undefined endogenous substrates, whereas the supply of defined substrates is an integral part of the original definition of State 3 and State 4, specifically achieved by depletion of undefined endogenous substrates in State 2 (Chance and Williams, 1955). (2) It is practically impossible to deplete an intact cell of ADP, but oligomycin is effective in living cells, such that a LEAK state can be induced with oligomycin (Lo).
- In the literature, State 4 is sometimes referred to as being equivalent to static head as defined in thermodynamics of irreversible processes. Thermodynamic steady states, however, do not refer to a specific protocol. Static head of mitochondrial respiration is a steady state maintaining a maximum output force (proton motive force or Gibbs force of phosphorylation) under a specific set of boundary conditions, among which the proton leak is a main (but not exclusive) component in determining the compensatory electron flux (measured as LEAK respiration). In contrast, State 4 describes a specific sequence in a defined protocol (Chance and Williams 1955), and this should not arbitrarily be used for entirely different protocols which may induce static head.
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