Template:SUIT-036

Steps and respiratory states

Step	State	Pathway	Q-junction	Comment - Events (E) and Marks (M)
1D	ROX			1D ADP is added to stimulate consumption of endogenous fuel-substrates. Rox is the residual oxygen consumption in the ROX state, due to oxidative side reactions, estimated either after inhibition of CIII (e.g. antimycin A, myxothiazol), CIV (e.g. Cyanide) or in the absence of endogenous fuel-substrates. Rox is subtracted from oxygen flux as a baseline for all respiratory states, to obtain mitochondrial respiration.
2M.1				1D;2M.1 Low concentration of malate, typically 0.1 mM, does not saturate the N-pathway; but saturates the F-pathway.
3Pal	PalMP	F	FAO	1D;2M.1;3Pal Respiratory stimulation of the FAO-pathway, F, by fatty acid, FA, in the presence of malate, M. Malate is a type N substrate (N), required for the F-pathway. The FA concentration has to be optimized to saturate the F-pathway, without inhibiting or uncoupling respiration.
3c	PalMcP	F	FAO	1D;2M.1;3Pal;3c Respiratory stimulation of the FAO-pathway, F, by fatty acid, FA, in the presence of malate, M. Malate is a type N substrate (N), required for the F-pathway. The FA concentration has to be optimized to saturate the F-pathway, without inhibiting or uncoupling 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]).
4M.2	PalMP	F(N)	FAO	1D;2M.1;3Pal;3c;4M.2 Respiratory stimulation of the FAO-pathway, F, by fatty acid FA in the presence of malate M. Malate is a type N substrate (N), required for the F-pathway. In the presence of anaplerotic pathways (e.g., mitochondrial malic enzyme, mtME) the F-pathway capacity is overestimated, if there is an added contribution of NADH-linked respiration, F(N) (see SUIT-002). The FA concentration has to be optimized to saturate the FAO-pathway, without inhibiting or uncoupling respiration. Malate kinetics in the presence of saturating [ADP] allows the evaluation of malate anaplerotic pathways. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
4M.5	PalMP	F(N)	FAO	1D;2M.1;3Pal;3c;4M.2;4M.5 Respiratory stimulation of the FAO-pathway, F, by fatty acid FA in the presence of malate M. Malate is a type N substrate (N), required for the F-pathway. In the presence of anaplerotic pathways (e.g., mitochondrial malic enzyme, mtME) the F-pathway capacity is overestimated, if there is an added contribution of NADH-linked respiration, F(N) (see SUIT-002). The FA concentration has to be optimized to saturate the FAO-pathway, without inhibiting or uncoupling respiration. Malate kinetics in the presence of saturating [ADP] allows the evaluation of malate anaplerotic pathways. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
4M1	PalMP	F(N)	FAO	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1 Respiratory stimulation of the FAO-pathway, F, by fatty acid FA in the presence of malate M. Malate is a type N substrate (N), required for the F-pathway. In the presence of anaplerotic pathways (e.g., mitochondrial malic enzyme, mtME) the F-pathway capacity is overestimated, if there is an added contribution of NADH-linked respiration, F(N) (see SUIT-002). The FA concentration has to be optimized to saturate the FAO-pathway, without inhibiting or uncoupling respiration. Malate kinetics in the presence of saturating [ADP] allows the evaluation of malate anaplerotic pathways. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
4M2	PalMP	F(N)	FAO	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2 Respiratory stimulation of the FAO-pathway, F, by fatty acid FA in the presence of malate M. Malate is a type N substrate (N), required for the F-pathway. In the presence of anaplerotic pathways (e.g., mitochondrial malic enzyme, mtME) the F-pathway capacity is overestimated, if there is an added contribution of NADH-linked respiration, F(N) (see SUIT-002). The FA concentration has to be optimized to saturate the FAO-pathway, without inhibiting or uncoupling respiration. Malate kinetics in the presence of saturating [ADP] allows the evaluation of malate anaplerotic pathways. High concentration of malate, typically 2 mM, saturates the N-pathway. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
5P	PalPMP	FN	FAO&CI	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P Respiratory stimulation by simultaneous action of the F-pathway and N-pathway with convergent electron flow in the FN-pathway for evaluation of an additive or inhibitory effect of F. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
6G	PalPGMP	FN	FAO&CI	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P;6G Respiratory stimulation by simultaneous action of the F-pathway and N-pathway with convergent electron flow in the FN-pathway for evaluation of an additive or inhibitory effect of F. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
7S10	PalPGMSP	FNS	FAO&CI&II	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P;6G;7S10 Respiratory stimulation by simultaneous action of the F-pathway, N-pathway, and S-pathway, with convergent electron flow in the FNS-pathway for reconstitution of TCA cycle function and additive or inhibitory effect of F. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
7S50	PalPGMSP	FNS	FAO&CI&II	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P;6G;7S10;7S50 Respiratory stimulation by simultaneous action of the F-pathway, N-pathway, and S-pathway, with convergent electron flow in the FNS-pathway for reconstitution of TCA cycle function and additive or inhibitory effect of F. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
8Gp	PalPGMSGpP	FNSGp	FAO&CI&II&GpDH	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P;6G;7S10;7S50;8Gp Respiratory stimulation by simultaneous action of fatty acid (F), type N substrates (N), succinate (S), and glycerophosphate with convergent electron flow in the FNSGp-pathway to the Q-junction. OXPHOS capacity P (with saturating [ADP]), active OXPHOS state.
9U	PalPGMSGpE	FNSGp	FAO&CI&II&GpDH	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P;6G;7S10;7S50;8Gp;9U Respiratory stimulation by simultaneous action of fatty acid (F), type N substrates (N), succinate (S), and glycerophosphate with convergent electron flow in the FNSGp-pathway to the Q-junction. 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.
10Rot	SGpE	SGp	CII&GpDH	1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P;6G;7S10;7S50;8Gp;9U;10Rot Respiratory stimulation by action of succinate and glycerophosphate, Gp, with convergent electron flow in the SGp-pathway (CII&GpDH-linked pathway to the Q-junction). Noncoupled electron transfer state, ET state, with ET capacity E.
11Ama	ROX			1D;2M.1;3Pal;3c;4M.2;4M.5;4M1;4M2;5P;6G;7S10;7S50;8Gp;9U;10Rot;11Ama 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).

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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