SUIT protocol library: Difference between revisions
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|abbr=SUITs | |abbr=SUITs | ||
|description=The '''Substrate-uncoupler-inhibitor titration (SUIT) protocol library''' contains classes of [[SUIT protocols]] with coupling and substrate control defined for [[mitochondrial preparations]]. | |description=The '''Substrate-uncoupler-inhibitor titration (SUIT) protocol library''' contains classes of [[SUIT protocols]] with coupling and substrate control defined for [[mitochondrial preparations]]. | ||
|info=[[MitoPedia: SUIT]] | |||
}} | }} | ||
{{MitoPedia concepts | {{MitoPedia concepts | ||
|mitopedia concept= | |mitopedia concept=MiP concept, SUIT concept | ||
}} | }} | ||
{{MitoPedia methods}} | {{MitoPedia methods}} | ||
{{MitoPedia O2k and high-resolution respirometry | {{MitoPedia O2k and high-resolution respirometry}} | ||
}} | |||
{{MitoPedia topics}} | {{MitoPedia topics}} | ||
'''''This page is in preparation''''' (intern wiki: [http://www.intern.oroboros.at/index.php/Protocol_development]) | '''''This page is in preparation''''' (intern wiki: [http://www.intern.oroboros.at/index.php/Protocol_development]) | ||
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== Strategy == | == Strategy == | ||
# Select a [[#Pathway control states |pathway control state]]. | :::# Select a [[#Pathway control states |pathway control state]]. | ||
# Select a [[#Control patterns |control pattern]]. | :::# Select a [[#Control patterns |control pattern]]. | ||
# [[#Define substrates |Define substrates]] for each pathway state. | :::# [[#Define substrates |Define substrates]] for each pathway state. | ||
# [[#Define coupling states |Define coupling states]]. | :::# [[#Define coupling states |Define coupling states]]. | ||
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=== Experimental conditions === | === Experimental conditions === | ||
* Temp | :::* Temp | ||
* O2 | :::* O2 | ||
* Medium | :::* Medium | ||
* Oxygen level | :::* Oxygen level | ||
* ADP kinetics | :::* ADP kinetics | ||
* Substrate kinetics | :::* Substrate kinetics | ||
=== O2k configuration === | === O2k configuration === | ||
* Amp: Amplex Red, TMRM, Ca Green, ... | :::* Amp: Amplex Red, TMRM, Ca Green, ... | ||
* Pot | :::* Pot | ||
* Spectrophotometry | :::* Spectrophotometry | ||
=== Define substrates === | === Define substrates === | ||
[[Substrate control state]]s are defined for each pathway control state ([[Complex I-linked substrate state |CI-linked]], [[Complex II-linked substrate state |CII-linked]], [[Complex I&II-linked substrate state |CI<small>&</small>II-linked]]) is defined by a specific substrate or substrate combination. These substrates may be added in a single step, or in multiple steps for analyis of the effect of different substrate combinations within a pathway control state. | :::: [[Substrate control state]]s are defined for each pathway control state ([[Complex I-linked substrate state |CI-linked]], [[Complex II-linked substrate state |CII-linked]], [[Complex I&II-linked substrate state |CI<small>&</small>II-linked]]) is defined by a specific substrate or substrate combination. These substrates may be added in a single step, or in multiple steps for analyis of the effect of different substrate combinations within a pathway control state. | ||
* CI: PM, GM, PGM, M, .. | Β | ||
* CII: S(Rot), S, SM, .. | :::* CI: PM, GM, PGM, M, .. | ||
* CI<small>&</small>II: PS, GS, PMS, GMS, PGMS, .. | :::* CII: S(Rot), S, SM, .. | ||
* CIV: AsTM | :::* CI<small>&</small>II: PS, GS, PMS, GMS, PGMS, .. | ||
* FAO: Oct, Pal, ... | :::* CIV: AsTM | ||
* CGpDH: Β | :::* FAO: Oct, Pal, ... | ||
* c | :::* CGpDH: Β | ||
:::* c | |||
=== Define coupling states === | === Define coupling states === | ||
The [[coupling control state]]s require a complementary definition of an [[ETS-competent substrate state]]. | :::: The [[coupling control state]]s require a complementary definition of an [[ETS-competent substrate state]]. | ||
* ''L'', ''P'', ''E'' ([[Uncoupler]]s, U: FCCP, CCCP, DNP) | :::* ''L'', ''P'', ''E'' ([[Uncoupler]]s, U: FCCP, CCCP, DNP) | ||
Coupling states: [[Image:L.jpg|link=LEAK respiration|LEAK]] [[Image:P.jpg|link=OXPHOS capacity|OXPHOS]] [[Image:E.jpg|link=ETS capacity|ETS]] | Coupling states: [[Image:L.jpg|link=LEAK respiration|LEAK]] [[Image:P.jpg|link=OXPHOS capacity|OXPHOS]] [[Image:E.jpg|link=ETS capacity|ETS]] | ||
=== Define residual oxygen consumption === | === Define residual oxygen consumption === | ||
* ROX: Rot&Ama, Rot&Mna&Ama, Myx, Cn, Az, .. | :::* ROX: Rot&Ama, Rot&Mna&Ama, Myx, Cn, Az, .. | ||
[[Image:ROX.jpg|link=Residual oxygen consumption|ROX]] | [[Image:ROX.jpg|link=Residual oxygen consumption|ROX]] | ||
== Categories of SUIT protocolsΒ == | == Categories of SUIT protocolsΒ == | ||
:Β» [[Categories of SUIT protocols]] | :::Β» [[Categories of SUIT protocols]] | ||
:Β» [[Talk:SUIT protocol library]] | :::Β» [[Talk:SUIT protocol library]] | ||
== SUIT protocol patterns == | == SUIT protocol patterns == | ||
:Β» [[SUIT protocol pattern]] | :::Β» [[SUIT protocol pattern]] | ||
== SUIT library == | == SUIT library == | ||
Β ''Work in progress'' | Β ''Work in progress'' | ||
:::Β» [[MitoPedia: SUIT]] | |||
=== CI + CI<small>&</small>II + CII diametral === | === CI + CI<small>&</small>II + CII diametral === | ||
Respiratory states: CI<sub>''L''</sub> -> CI<sub>''P''</sub> -> CI<small>&</small>II<sub>''P''</sub> -> CI<small>&</small>II<sub>''E''</sub> -> CII<sub>''E''</sub> -> ROX | :::: Respiratory states: CI<sub>''L''</sub> -> CI<sub>''P''</sub> -> CI<small>&</small>II<sub>''P''</sub> -> CI<small>&</small>II<sub>''E''</sub> -> CII<sub>''E''</sub> -> ROX | ||
* '''GM+D+S+U+Rot''' ''Innsbruck-Copenhagen protocol'': GM+D+c+S+U+Rot ::Cyt c: added after ADP; cyt c added after S; ADP conc: _ _; ROX: Ama ([[Boushel 2007 Diabetologia]]) | ::::* '''GM+D+S+U+Rot''' ''Innsbruck-Copenhagen protocol'': GM+D+c+S+U+Rot ::Cyt c: added after ADP; cyt c added after S; ADP conc: _ _; ROX: Ama ([[Boushel 2007 Diabetologia]]) | ||
* '''PMG+D+S+U+Rot''' ''Brain protocol'': PMG+D+c+S+U+Rot::ADP conc: in mouse brain homogenate a final conc. of 2 mM ADP in the [[OXPHOS]] state (CI<small>&</small>II) is saturating ([[Sumbalova 2011 Abstract Bordeaux]]); [[ROX]]: Mna&Ama; Myx | ::::* '''PMG+D+S+U+Rot''' ''Brain protocol'': PMG+D+c+S+U+Rot::ADP conc: in mouse brain homogenate a final conc. of 2 mM ADP in the [[OXPHOS]] state (CI<small>&</small>II) is saturating ([[Sumbalova 2011 Abstract Bordeaux]]); [[ROX]]: Mna&Ama; Myx | ||
=== CI + CI + CI<small>&</small>II + CII diamteral === | === CI + CI + CI<small>&</small>II + CII diamteral === | ||
* '''GM+D+P+S+U+Rot''' ''Innsbruck-Liege protocol 1'': GM+D+c+P+S+U+Rot::[[Cytochrome c]]: early addition of ''c'' ensures comparability of all states in case of any ''c'' effect. | ::::* '''GM+D+P+S+U+Rot''' ''Innsbruck-Liege protocol 1'': GM+D+c+P+S+U+Rot::[[Cytochrome c]]: early addition of ''c'' ensures comparability of all states in case of any ''c'' effect. | ||
* '''PM+D+G+S+U+Rot''' ''Innsbruck-Liege protocol 2'': PM+D+c+G+S+U+Rot Β | ::::* '''PM+D+G+S+U+Rot''' ''Innsbruck-Liege protocol 2'': PM+D+c+G+S+U+Rot Β | ||
=== FAO + CI + CI<small>&</small>II + CII diametral === | === FAO + CI + CI<small>&</small>II + CII diametral === | ||
Respiratory states: FAO<sub>''L''</sub> -> FAO<sub>''P''</sub> -> CI<small>&</small>FAO<sub>''P''</sub> -> CI<small>&</small>II<small>&</small>FAO<sub>''P''</sub> -> CI<small>&</small>II<small>&</small>FAO<sub>''E''</sub> -> CII<sub>''E''</sub> -> ROX | :::: Respiratory states: FAO<sub>''L''</sub> -> FAO<sub>''P''</sub> -> CI<small>&</small>FAO<sub>''P''</sub> -> CI<small>&</small>II<small>&</small>FAO<sub>''P''</sub> -> CI<small>&</small>II<small>&</small>FAO<sub>''E''</sub> -> CII<sub>''E''</sub> -> ROX | ||
* '''OctM+D+G+S+U+Rot''' ''Innsbruck training protocol'': OctM+D+G+c+S+U+Rot::[[Pesta 2012 Methods Mol Biol]], [[Pesta 2011 Am J Physiol Regul Integr Comp Physiol]] | ::::* '''OctM+D+G+S+U+Rot''' ''Innsbruck training protocol'': OctM+D+G+c+S+U+Rot::[[Pesta 2012 Methods Mol Biol]], [[Pesta 2011 Am J Physiol Regul Integr Comp Physiol]] | ||
Respiratory states: FAO<sub>''L''</sub> -> FAO<sub>''P''</sub> -> CI<small>&</small>FAO<sub>''P''</sub> -> CI<small>&</small>II<small>&</small>FAO<sub>''P''</sub> -> CII<sub>''P''</sub> -> CII<sub>''L''</sub> -> CII<sub>''E''</sub> -> ROX | :::: Respiratory states: FAO<sub>''L''</sub> -> FAO<sub>''P''</sub> -> CI<small>&</small>FAO<sub>''P''</sub> -> CI<small>&</small>II<small>&</small>FAO<sub>''P''</sub> -> CII<sub>''P''</sub> -> CII<sub>''L''</sub> -> CII<sub>''E''</sub> -> ROX | ||
* '''OctM+D+G+S+Rot+Omy+U''' ''Greenland-Monte Rosa protocol'': OctM+D+G+S+Rot+Omy+U+c::[[Cytochrome c|Cyt ''c'']]: added after U. The absence of a significant ''c''-effect shows the integrity of the outer mitochondrial membrane after fibre preparation and also shows the preservation of integrity over a 90-100 min incubation in the O2k ([[MiR05]]); Oxygen: oxygen limitation of flux was preventd by maintaining oxygen levels in the respirometer above air saturation ([[Pesta 2012 Methods Mol Biol]]); [[ROX]]: Mna+Ama | ::::* '''OctM+D+G+S+Rot+Omy+U''' ''Greenland-Monte Rosa protocol'': OctM+D+G+S+Rot+Omy+U+c::[[Cytochrome c|Cyt ''c'']]: added after U. The absence of a significant ''c''-effect shows the integrity of the outer mitochondrial membrane after fibre preparation and also shows the preservation of integrity over a 90-100 min incubation in the O2k ([[MiR05]]); Oxygen: oxygen limitation of flux was preventd by maintaining oxygen levels in the respirometer above air saturation ([[Pesta 2012 Methods Mol Biol]]); [[ROX]]: Mna+Ama | ||
== References and notes == | == References and notes == | ||
* [[MitoPedia: Respiratory states]] | :::* [[MitoPedia: Respiratory states]] | ||
* [[MitoPedia: Terms and abbreviations]] | :::* [[MitoPedia: Terms and abbreviations]] | ||
* [[Gnaiger_2014_MitoPathways]] | ::::* [[Gnaiger_2014_MitoPathways]] | ||
* [[Gnaiger_2009_Int J Biochem Cell Biol]] | ::::* [[Gnaiger_2009_Int J Biochem Cell Biol]] | ||
* [[Pesta 2012 Methods Mol Biol]] | ::::* [[Pesta 2012 Methods Mol Biol]] | ||
* [[Pesta 2011 Am J Physiol Regul Integr Comp Physiol]] | ::::* [[Pesta 2011 Am J Physiol Regul Integr Comp Physiol]] | ||
* [[Sumbalova 2011 Abstract Bordeaux]] | ::::* [[Sumbalova 2011 Abstract Bordeaux]] |
Revision as of 09:05, 15 May 2016
Description
The Substrate-uncoupler-inhibitor titration (SUIT) protocol library contains classes of SUIT protocols with coupling and substrate control defined for mitochondrial preparations.
Abbreviation: SUITs
Reference: MitoPedia: SUIT
MitoPedia concepts:
MiP concept,
SUIT concept
This page is in preparation (intern wiki: [1])
Strategy
- Select a pathway control state.
- Select a control pattern.
- Define substrates for each pathway state.
- Define coupling states.
Definition of states
Experimental conditions
- Temp
- O2
- Medium
- Oxygen level
- ADP kinetics
- Substrate kinetics
O2k configuration
- Amp: Amplex Red, TMRM, Ca Green, ...
- Pot
- Spectrophotometry
Define substrates
- Substrate control states are defined for each pathway control state (CI-linked, CII-linked, CI&II-linked) is defined by a specific substrate or substrate combination. These substrates may be added in a single step, or in multiple steps for analyis of the effect of different substrate combinations within a pathway control state.
- CI: PM, GM, PGM, M, ..
- CII: S(Rot), S, SM, ..
- CI&II: PS, GS, PMS, GMS, PGMS, ..
- CIV: AsTM
- FAO: Oct, Pal, ...
- CGpDH:
- c
Define coupling states
- The coupling control states require a complementary definition of an ETS-competent substrate state.
- L, P, E (Uncouplers, U: FCCP, CCCP, DNP)
Define residual oxygen consumption
- ROX: Rot&Ama, Rot&Mna&Ama, Myx, Cn, Az, ..
Categories of SUIT protocols
SUIT protocol patterns
SUIT library
Work in progress
CI + CI&II + CII diametral
- Respiratory states: CIL -> CIP -> CI&IIP -> CI&IIE -> CIIE -> ROX
- GM+D+S+U+Rot Innsbruck-Copenhagen protocol: GM+D+c+S+U+Rot ::Cyt c: added after ADP; cyt c added after S; ADP conc: _ _; ROX: Ama (Boushel 2007 Diabetologia)
- PMG+D+S+U+Rot Brain protocol: PMG+D+c+S+U+Rot::ADP conc: in mouse brain homogenate a final conc. of 2 mM ADP in the OXPHOS state (CI&II) is saturating (Sumbalova 2011 Abstract Bordeaux); ROX: Mna&Ama; Myx
CI + CI + CI&II + CII diamteral
- GM+D+P+S+U+Rot Innsbruck-Liege protocol 1: GM+D+c+P+S+U+Rot::Cytochrome c: early addition of c ensures comparability of all states in case of any c effect.
- PM+D+G+S+U+Rot Innsbruck-Liege protocol 2: PM+D+c+G+S+U+Rot
FAO + CI + CI&II + CII diametral
- Respiratory states: FAOL -> FAOP -> CI&FAOP -> CI&II&FAOP -> CI&II&FAOE -> CIIE -> ROX
- OctM+D+G+S+U+Rot Innsbruck training protocol: OctM+D+G+c+S+U+Rot::Pesta 2012 Methods Mol Biol, Pesta 2011 Am J Physiol Regul Integr Comp Physiol
- Respiratory states: FAOL -> FAOP -> CI&FAOP -> CI&II&FAOP -> CIIP -> CIIL -> CIIE -> ROX
- OctM+D+G+S+Rot+Omy+U Greenland-Monte Rosa protocol: OctM+D+G+S+Rot+Omy+U+c::Cyt c: added after U. The absence of a significant c-effect shows the integrity of the outer mitochondrial membrane after fibre preparation and also shows the preservation of integrity over a 90-100 min incubation in the O2k (MiR05); Oxygen: oxygen limitation of flux was preventd by maintaining oxygen levels in the respirometer above air saturation (Pesta 2012 Methods Mol Biol); ROX: Mna+Ama