Template:SUIT-006 General O2 pce D053

Steps and respiratory states

Step	State	Pathway	Q-junction	Comment - Events (E) and Marks (M)
ce1	ROUTINE			ce1 ROUTINE respiration in the physiological coupling state R. Externally added permeable substrates could contribute to this respiratory state.
1Dig	REN			ce1;1Dig Optimum effective digitonin concentration for complete plasma membrane permeabilization.

Step	State	Pathway	Q-junction	Comment - Events (E) and Marks (M)
1X	XL			ce1;Dig1;1X Electron-transfer-pathway state will depend on the substrate/inhibitor combinations added to fuel the mitochondrial respiration Non-phosphorylating resting state (LEAK state); LEAK respiration L(n) in the absence of ADP, ATP, AMP (no adenylates).
2D	XP			ce1;Dig1;1X;2D Electron-transfer-pathway state will depend on the substrate/inhibitor combinations added to fuel the mitochondrial respiration OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
2c	XcP			ce1;Dig1;1X;2D;2c Electron-transfer-pathway state will depend on the substrate/inhibitor combinations added to fuel the mitochondrial respiration OXPHOS capacity P (with saturating [ADP]), active OXPHOS state. Addition of cytochrome c yields a test for integrity of the mtOM (cytochrome c control efficiency). Stimulation by added cytochrome c would indicate an injury of the mtOM and limitation of respiration in the preceding state without added c due to loss of cytochrome c. Typically, cytochrome c is added immediately after the earliest ADP-activation step (OXPHOS capacity P with saturating [ADP]).
3(Omy)	XL			ce1;Dig1;1X;2D;2c;3(Omy) Non-phosphorylating resting state (LEAK state); LEAK-respiration, L(Omy), after blocking the ATP synthase with oligomycin. The addition of oligomycin or its carrier (EtOH) is optional depending on the experimental question to resolve. If we add only the carrier we do not reach LEAK state and we will evaluate if the EtOH alters the ROUTINE state.
4U	XE			ce1;Dig1;1X;2D;2c;3(Omy);4U Electron-transfer-pathway state will depend on the substrate/inhibitor combinations added to fuel the mitochondrial respiration Uncoupler titration (avoiding inhibition by high uncoupler concentrations) to obtain electron transfer (ET) capacity E (noncoupled ET-state). Test for limitation of OXPHOS capacity P by the phosphorylation system (ANT, ATP synthase, phosphate transporter) relative to ET capacity E in mt-preparations: E-P control efficiency and E-L coupling efficiency. In living cells: E-R control efficiency and E-L coupling efficiency.
5*	ROX			ce1;Dig1;1X;2D;2c;3(Omy);4U;5* (*) Rox obtaining after inhibiting the CIII (e.g. antimycin A, myxothiazol). Rox is the residual oxygen consumption in the ROX state, due to oxidative side reactions, estimated after addition of antimycin A (inhibitor of CIII). Rox is subtracted from oxygen flux as a baseline for all respiratory states, to obtain mitochondrial respiration (mt).

Step	Respiratory state	Pathway control	ET-Complex	Comment
## AsTm	AsTmE	CIV	CIV
## Azd	CHB

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Coupling control

» Coupling control state

» ET capacity

» OXPHOS capacity

» LEAK respiration

Pathway control

» Electron transfer pathway

» Fatty acid oxidation pathway control state, F

» NADH electron transfer-pathway state, N

» Succinate pathway control state, S

» NS-pathway control state, NS

» Glycerophosphate pathway control state, Gp

» Complex IV single step, CIV

» Anaplerotic pathway control state

Main fuel substrates

» Glutamate, G

» Glycerophosphate, Gp

» Malate, M

» Octanoylcarnitine, Oct

» Pyruvate, P

» Succinate, S

Glossary

» List of SUIT states

» SUIT concept