From Bioblast
Description
Abbreviation: N(PM)
Reference: A SUIT-006 is to analyse the coupling control in NADH-pathway on mitochondrial prepartions (isolated mitochondria and tissue homogenate).
SUIT number: D047_1PM;2D;2c;3Omy;4U;5Ama
O2k-Application: O2
- MitoPedia: SUIT
- SUIT-category: N(PM)
- SUIT protocol pattern: Bidirectional linear coupling control protocol 1PM;2D;2c;3Omy;4U;5Ama
SUIT-006 O2 mt D047 is a coupling control protocol for for mitochondrial preparations (tissue homogenate and isolated mitochondria). Different coupling control states are assessed (L(n) - P - E or L(n) - P - L(Omy) - E) at N(PM) pathway control state. The use of oligomycin is optional, however, it provides important information when ATPases are present, a situation that may lead to overestimated L(n) respiration.
Communicated by Cardoso LH, Komlodi T and Gnaiger E (last update 2019-03-05)
Steps and respiratory states
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M)
|
---|---|---|---|---|
1PM | PML(n) | N | CI | 1PM
|
2D | PMP | N | CI | 1PM;2D
|
2c | PMcP | N | CI | 1PM;2D;2c
|
3Omy | PML(Omy) | N | CI | 1PM;2D;2c;3Omy
|
4U | PME | N | CI | 1PM;2D;2c;3Omy;4U
|
5Ama | ROX | 1PM;2D;2c;3Omy;4U;5Ama
|
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- Coupling control
- Pathway control
- Β» Electron transfer pathway
- Β» Fatty acid oxidation pathway control state, F
- Β» NADH electron transfer-pathway state, N
- Β» Succinate pathway control state, S
- Β» NS-pathway control state, NS
- Β» Glycerophosphate pathway control state, Gp
- Β» Complex IV single step, CIV
- Β» Anaplerotic pathway control state
- Pathway control
- Main fuel substrates
- Β» Glutamate, G
- Β» Glycerophosphate, Gp
- Β» Malate, M
- Β» Octanoylcarnitine, Oct
- Β» Pyruvate, P
- Β» Succinate, S
- Main fuel substrates
- Glossary
Strengths and limitations
- This protocol is indicated when the aim is to analyse coupling control in isolated mitochondria or tissue homogenates in NADH-pathway using PM as substrates.
- + Reasonable duration of the experiment.
- + This protocol can be extended with the Complex IV module.
- - This protocol analyses the coupling control in NADH-pathway (with PM), however, it is less specific than a similar protocol containing the complex II inhibitor malonic acid, that would allow to analyse the NADH-pathway without any contribution of the S-pathway.
- - Careful washing is required after the experiment to avoid carry-over of uncoupler and inhibitors, if used.
Compare SUIT protocols
- SUIT-001 O2 mt D001: Extended protocol with SUIT-006 without oligomycin as the initial coupling control module, for mitochondrial preparations.
- SUIT-003: Coupling control protocol for non-permeabilized cells (ce), counterpart to SUIT-006 for mt-preparations (mtprep).
References
MitoPedia concepts: SUIT protocol, SUIT A, Find
MitoPedia methods:
Respirometry