Difference between revisions of "SUIT-012 O2 ce-pce D052"
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{{MitoPedia | {{MitoPedia | ||
|abbr=N(PGM) | |abbr=N(PGM) | ||
|description=[[File:ce1;1Dig;1PM;2D;2c;3G;4U;5Ama.png| | |description=[[File:ce1;1Dig;1PM;2D;2c;3G;4U;5Ama.png|450px]] | ||
|info='''A''' [[SUIT-012]] | |info='''A''' Linear coupling control with NADH-linked substrates for pce - '''[[SUIT-012]]''' | ||
|application=O2 | |application=O2 | ||
|SUIT number= | |SUIT number=D052_ce1;1Dig;1PM;2D;2c;3G;4U;5Ama | ||
}} | }} | ||
::: '''[[Categories of SUIT protocols |SUIT-category]]:''' N(PGM) | ::: '''[[Categories of SUIT protocols |SUIT-category]]:''' N(PGM) | ||
::: '''[[SUIT protocol pattern]]:''' diametral ce1;1Dig;1PM;2D;2c;3G;4U | ::: '''[[SUIT protocol pattern]]:''' diametral ce1;1Dig;1PM;2D;2c;3G;4U- | ||
{{Template:SUIT text D052}} | |||
__TOC__ | __TOC__ | ||
Communicated by [[Huete-Ortega M]], [[Komlodi T]], [[Gnaiger E]] (last update 2019-04-03) | |||
== Steps and respiratory States == | |||
== Representative traces == | |||
[[File:D052 O2 traces.png|600px]] | |||
{{Template:SUIT-012 O2 ce-pce D052}} | |||
== Strengths and limitations == | |||
:::+ This protocol allows to evaluate the function of the TCA cycle without the involvement of the [[complex II]] (S-pathway). Would be useful to understand the contribution and the activity of the dehydrogenases. | |||
:::+ Mitochondrial outer membrane integrity can be measured with the addition of [[cytochrome c]]. Application of the [[cytochrome c]] test early in the protocol ensures comparability of all states in case of any effect of cytochrome c. | |||
:::+ Reasonable duration of the experiment. | |||
:::+ This protocol can be extended with the Complex IV module, which can prolong the experimental time with ~30 min. | |||
:::+ GM and PM yield typically identical fluxes in human skeletal muscle fibres. However, PM is the superior alternative to GM: the fraction of the N-pathway is lower and of the S-pathway is higher with GM compared to PM (GMP is inhibited by the CII inhibitor malonic acid to a larger extent than PMP). PM, therefore, yields a more sensitive assay for the diagnosis of injuries in the N-pathway, since an impairment of N-pathway capacity can be compensated partially by activation of the S-pathway. This is an advantage compared to [[SUIT-011]] for diagnosis of N-capacity. | |||
:::- Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler. | |||
== Compare SUIT protocols == | |||
::::* [[SUIT-008]]: 1PM;2D;3G;4S;5U-; longer version that also analyses S-pathway. | |||
::::* [[SUIT-006]]: 1X;2D;3Omy;4U-; coupling control protocol with only one combination of substrates (''e.g.'' 1PM;2D;3Omy;4U-). | |||
== Chemicals and syringes == | |||
{{Template:Chemicals ce}} | |||
{{Template:Chemicals SUIT-012}} | |||
::: Suggested stock concentrations are shown in the specific DL-Protocol. | |||
== References == | == References == | ||
{{#ask:[[Category:Publications]] [[Instrument and method::O2k-Protocol]] [[Additional label::SUIT-012]] | {{#ask:[[Category:Publications]] [[Instrument and method::O2k-Protocol]] [[Additional label::SUIT-012 ce-pce D052]] | ||
|?Was published in year=Year | |?Was published in year=Year | ||
|?Has title=Reference | |?Has title=Reference | ||
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|order=descending | |order=descending | ||
}} | }} | ||
{{MitoPedia concepts | {{MitoPedia concepts | ||
|mitopedia concept=SUIT protocol, SUIT A, Find | |mitopedia concept=SUIT protocol, SUIT A, Find |
Latest revision as of 00:14, 13 April 2021
Description
Abbreviation: N(PGM)
Reference: A Linear coupling control with NADH-linked substrates for pce - SUIT-012
SUIT number: D052_ce1;1Dig;1PM;2D;2c;3G;4U;5Ama
O2k-Application: O2
- SUIT-category: N(PGM)
- SUIT protocol pattern: diametral ce1;1Dig;1PM;2D;2c;3G;4U-
The SUIT-012 O2 ce-pce D052 protocol specifically focuses on assessing the coupling control (L- P- E) with NADH-linked substrates (PM and PGM). Addition of G enables evaluating the contribution of this substrate in NADH-supported OXPHOS respiration, which may be relevant in the presence of glutamate-anaplerotic pathway control state. SUIT-012 can be extended with the CIV assay module.
Communicated by Huete-Ortega M, Komlodi T, Gnaiger E (last update 2019-04-03)
Steps and respiratory States
Representative traces
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M) |
---|---|---|---|---|
ce1 | ROUTINE | ce1
| ||
1Dig | REN | ce1;1Dig
|
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
|
3G | PGMP | N | CI | 1PM;2D;2c;3G
|
4U | PGME | N | CI | 1PM;2D;2c;3G;4U
|
5Ama | ROX | 1PM;2D;2c;3G;4U;5Ama
|
Step | Respiratory state | Pathway control | ET-Complex | Comment |
---|---|---|---|---|
## AsTm | AsTmE | CIV | CIV | |
## Azd | CHB |
- Bioblast links: SUIT protocols - >>>>>>> - Click on [Expand] or [Collapse] - >>>>>>>
- Coupling control
- Pathway control
- Main fuel substrates
- » Glutamate, G
- » Glycerophosphate, Gp
- » Malate, M
- » Octanoylcarnitine, Oct
- » Pyruvate, P
- » Succinate, S
- Main fuel substrates
- Glossary
Strengths and limitations
- + This protocol allows to evaluate the function of the TCA cycle without the involvement of the complex II (S-pathway). Would be useful to understand the contribution and the activity of the dehydrogenases.
- + Mitochondrial outer membrane integrity can be measured with the addition of cytochrome c. Application of the cytochrome c test early in the protocol ensures comparability of all states in case of any effect of cytochrome c.
- + Reasonable duration of the experiment.
- + This protocol can be extended with the Complex IV module, which can prolong the experimental time with ~30 min.
- + GM and PM yield typically identical fluxes in human skeletal muscle fibres. However, PM is the superior alternative to GM: the fraction of the N-pathway is lower and of the S-pathway is higher with GM compared to PM (GMP is inhibited by the CII inhibitor malonic acid to a larger extent than PMP). PM, therefore, yields a more sensitive assay for the diagnosis of injuries in the N-pathway, since an impairment of N-pathway capacity can be compensated partially by activation of the S-pathway. This is an advantage compared to SUIT-011 for diagnosis of N-capacity.
- - Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler.
Compare SUIT protocols
Chemicals and syringes
Step | Chemical(s) and link(s) | Comments |
---|---|---|
1Dig | Digitonin (Dig) |
Step | Chemical(s) and link(s) | Comments |
---|---|---|
1PM | Pyruvate (P) and Malate (M) | |
2D | ADP (D) | |
2c | Cytochrome c (c) | |
3G | Glutamate (G) | |
4U | Carbonyl cyanide m-chlorophenyl hydrazone, CCCP (U) | Can be substituted for other uncoupler |
5Ama | Antimycin A (Ama) |
- Suggested stock concentrations are shown in the specific DL-Protocol.
References
MitoPedia concepts: SUIT protocol, SUIT A, Find
MitoPedia methods:
Respirometry