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Difference between revisions of "SUIT protocol library"

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=== 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''
* ''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, 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]]


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# '''Diametral coupling -> substrate ->> control''' (mono- or bidirectional)
# '''Diametral coupling -> substrate ->> control''' (mono- or bidirectional)
# '''Diametral substrate -> coupling ->> control''' (mono- or bidirectional)
# '''Diametral substrate -> coupling ->> control''' (mono- or bidirectional)
== SUIT library ==
=== CI + CI<small>&</small>CII + 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
* '''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
=== CI + CI + CI<small>&</small>CII + 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<small>&</small>CII + 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
* '''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
* '''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




Revision as of 04:26, 19 August 2015

This page is in preparation (intern wiki: SUIT protocol library intern)

Substrate-uncoupler-inhibitor titration (SUIT) protocols

Classes of SUIT protocols with coupling and substrate control defined for mitochondrial preparations


Strategy

  1. Select a pathway control state.
  2. Select a control pattern.
  3. Define substrates for each pathway state.
  4. Define coupling states.


Definition of states

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.

Coupling states: LEAK OXPHOS ETS


Define residual oxygen consumption

  • ROX: Rot&Ama, Rot&Mna&Ama, Myx, Cn, Az, ..

ROX


Pathway control states

Single pathway control states

  1. CI
  2. CII
  3. CI&CII
  4. CIV
  5. FAO
  6. CGPDH

Multiple pathway control states

A. CI-CII-CIV

  1. CI + CII
  2. CI + CII + CIV
  3. CI + CI&CII + CII
  4. CI + CI&CII + CII + CIV

B. FAO-CI-CII-CIV

  1. FAO + CI&FAO + CII
  2. FAO + CI&FAO + CII + CIV
  3. FAO + CI&FAO + CI&CII&FAO + CII
  4. FAO + CI&FAO + CI&CII&FAO + CII + CIV

C. CGPDH-CI-CII-CIV

  1. CGPDH + CI&CGPDH
  2. CGPDH + CI&CGPDH + CII&CGPDH
  3. CGPDH + CI&CGPDH + CII&CGPDH + CIV
  4. CGPDH + CI&CGPDH + CI&CII&CGPDH + CII&CGPDH
  5. CGPDH + CI&CGPDH + CI&CII&CGPDH + CII&CGPDH + CIV
  6. CI + CI&CGPDH + ...
  7. CI + CI&CII + CI&CII&CGPDH + ...
  8. CI&CII + CI&CII&CGPDH + ...

D. FAO-CGPDH-CI-CII-CIV

  1. FAO + FAO&CGPDH + CI&FAO&CGPDH + ...
  2. CGPDH + FAO&CGPDH + CI&FAO&CGPDH + ...
  3. CI + CI&FAO + CI&FAO&CGPDH + ...
  4. CI + CI&CGPDH + CI&FAO&CGPDH + ...
  5. ...


Control patterns

Linear

  1. Linear coupling control: at a single substrate state (classical State 3 -> State 4, or P -> LT).
  2. Linear substrate control: at a single coupling state (unusual).
  3. Monodirectional linear coupling control: LN -> PD -> EF
  4. Bidirectional linear coupling control: LN -> PD -> LOmy -> EF


Orthogonal

  1. Orthogonal coupling -> substrate control (mono- or bidirectional)
  2. Orthogonal substrate -> coupling control (mono- or bidirectional)


Diametral

  1. Diametral coupling -> substrate ->> control (mono- or bidirectional)
  2. Diametral substrate -> coupling ->> control (mono- or bidirectional)


SUIT library

CI + CI&CII + 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)


CI + CI + CI&CII + 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&CII + CII diametral

Respiratory states: FAOL -> FAOP -> CI&FAOP -> CI&II&FAOP -> CI&II&FAOE -> CIIE -> ROX


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


References and notes

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