The protonmotive force and respiratory control: Difference between revisions

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{{MITOEAGLE}}
{{MITOEAGLE}}
[[File:OXPHOS system.jpg|right|400px|thumb|Fig. 1. The mitochondrial respiratory system. In oxidative phosphorylation the electron transfer system (A) is coupled to the phosphorylation system (B). See Eqs. 4 and 5 for further explanation.]]
::: <big>'''Building blocks of mitochondrial physiology'''</big> '''MitoEAGLE recommendations Part 1.'''
<br />


== Mitochondrial respiratory control: a conceptual perspective on coupling states in mitochondrial preparations ==
::: '''MitoEAGLE recommendations Part 1.'''


:::» [[Talk:Mitochondrial respiratory control: MitoEAGLE recommendations |''Work in progress'']]: '''Download last update (Version 19) 2017-08-19''': [[File:PDF.jpg|120px|link=http://www.mitoglobal.org/images/0/01/MitoEAGLE_1_Mitochondrial_respiratory_coupling_control.pdf |Bioblast pdf]] - » [http://www.mitoeagle.org/index.php/File:MitoEAGLE_1_Mitochondrial_respiratory_coupling_control.pdf Versions]
{{MitoEAGLE preprint 1 Phases}}
[[File:Coupling in OXPHOS.jpg|right|400px|thumb|Fig. 2. The proton circuit and coupling in oxidative phosphorylation (OXPHOS). Modified after [[Gnaiger_2014_MitoPathways#Chapter_1._Real-time_OXPHOS_analysis |Gnaiger 2014 MitoPathways]].]]




::: '''MitoEAGLE Terminology Group'''
== Open invitation ==
::::* First draft, updates, and '''corresponding author''': [[Gnaiger E |E. Gnaiger]]
::::* '''Contributing co-authors''': ''Confirming to have read the final manuscript, possibly to have made additions or suggestions for improvement, and to agree to implement the recommendations into future manuscripts, presentations and teaching materials. (alphabetical, to be extended)'':
::::::* M.G. Alves, D. Ben-Shachar, G.C. Brown, G.R. Buettner, E. Calabria, A.J. Chicco, P.M. Coen, J.L. Collins, L. Crisóstomo, M.S. Davis, C. Doerrier, E. Elmer, A. Filipovska, P.M. Garcia-Roves, D.K. Harrison, K.T. Hellgren, C.L. Hoppel, J. Iglesias-Gonzalez, P. Jansen-Dürr, B.H. Goodpaster, B.A. Irving, S. Iyer, T. Komlodi, V. Laner, H.K. Lee, H. Lemieux, A.T. Meszaros, N. Moisoi, A. Molina, A.L. Moore, A.J. Murray, J. Neuzil, R.K. Porter, K. Nozickova, P.J. Oliveira, K. Renner-Sattler, J. Rohlena, D. Salvadego, L.A. Sazanov, O. Sobotka, R. Stocker, I. Szabo, M. Tanaka, L. Tretter, B. Velika, A.E. Vercesi, Y.H. Wei


::::* '''Supporting co-authors''': ''Confirming to have read the final manuscript, and to agree to implement the recommendations into future manuscripts, presentations and teaching materials. (alphabetical, to be extended)'':
:::: ''This manuscript on ‘The protonmotive force and respiratory control’ is a position statement in the frame of COST Action CA15203 MitoEAGLE. The list of coauthors evolved from MitoEAGLE Working Group Meetings and a bottom-up spirit of COST: This is an open invitation to scientists and students to join as coauthors, to provide a balanced view on mitochondrial respiratory control, a fundamental introductory presentation of the concept of the protonmotive force, and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. We plan a series of follow-up reports by the expanding MitoEAGLE Network, to increase the scope of consensus-oriented recommendations and facilitate global communication and collaboration.''
::::::* R.A. Brown, T. Dias, G. Distefano, H. Dubouchaud, L.F. Garcia-Souza, T. Käämbre, G. Keppner, A. Krajcova, M. Markova, J. Muntané, D. O'Gorman, M.T. Oliveira, C.M. Palmeira, P.X. Petit, K. Siewiera, P. Stankova, Z. Sumbalova


::::* '''Journal''': Int J Biochem Cell Biol (as discussed at [[MITOEAGLE Barcelona 2017]]); Open Access is a requirement. Final decision has to be made.
::::* ''We continue to invite comments and suggestions on the MitoEAGLE preprint (phase 2; until '''October 12'''), particularly if you are an early career investigator adding an open future-oriented perspective, or an established scientist providing a balanced historical basis. Your critical input into the quality of the manuscript will be most welcome, improving our aims to be educational, general, consensus-oriented, and practically helpful for students working in mitochondrial respiratory physiology.''
<br />


::: <big><big>'''Contents'''</big></big>
::::* '''''Please feel free to focus on a particular section in terms of direct input and references, while evaluating the entire scope of the manuscript from the perspective of your expertise.'''''


:::: '''1. Introduction'''
::::* Your comments will be largely posted on the discussion page of the MitoEAGLE preprint website. If you prefer to submit comments in the format of a referee's evaluation rather than a contribution as a coauthor, I will be glad to distribute your views to the updated list of coauthors for a balanced response. We would ask for your consent on this open bottom-up procedure.  
:::::::: Mitochondrial preparations, mtprep,
::: '''2. Fundamental respiratory coupling states in mitochondrial preparations'''
:::::: ''2.1. Definitions''
:::::::: Phosphorylation, »P
:::::::: Control and regulation
:::::: ''2.2. Classical terminology for isolated mitochondria''
:::::::: States 1-5
:::::: ''2.3. Three coupling states of mitochondrial preparations and residual oxygen consumption''
:::::::: OXPHOS, ETS, LEAK, ROX
::: '''3. States and rates'''
:::::: ''3.1. Respiratory states and respiratory rates''
:::::: ''3.2. The steady-state and protonmotive force''
:::::::: Protonmotive force, ∆''p''<sub>mt</sub>
:::::::: Faraday constant, ''F''
:::::::: Electrical part of the protonmotive force, el
:::::::: Chemical diffusion (or dislocation) part of the protonmotive force, d
:::::::: ''e''-isomorph [C]
:::::::: ''n''-isomorph [mol]
:::::: ''3.3. Forces and flows in physics and irreversible thermodynamics''
:::::::: Molar quantities
:::::::: Vectorial and scalar forces and fluxes
:::::::: Coupled versus bound processes
:::::::: Coupling, efficiency and power
:::::: ''3.4. Normalization: flows and fluxes''
:::::::: Extensive quantities
:::::::: Size-specific quantities
:::::::: Flow per system, ''I''
:::::::: Size-specific flux, ''J''
:::::::: Flux per volume of the instrumental system, ''J<sub>V</sub>''
:::::::: Flow per experimental model, ''I''
:::::: ''3.5. Conversion: oxygen, protons, ATP
::: '''4. Conclusions'''


New Fig. 6 and text in Version 16
:::: ''We organize a MitoEAGLE session linked to our series of communications at the MiP''conference'' Nov 2017 in Hradec Kralove in close association with the MiP''society'' (where you hopefully will attend) and at EBEC 2018 in Budapest.''
[[File:OXPHOS compartments.jpg|right|400px|thumb|Fig. 6. Four-compartmental model of oxidative phosphorylation with respiratory states (ETS, OXPHOS, LEAK) and corresponding rates (''E, P, L''). Modified from Gnaiger (2014).]]
:::::::::::: » '''[[MiP2017_Hradec_Kralove_CZ]]'''  
* 2018-08-18 Erich Gnaiger edited some equations in Versin 18:
:::: '''Protonmotive force, ∆''p''<sub>mt</sub>''': The protonmotive force, ∆''p''<sub>mt</sub>,
:::::::::::::: ∆''p''<sub>mt</sub> = ∆''Ψ''<sub>mt</sub> + ∆''µ''<sub>H+</sub> / ''F'' ; (Eq. 1)
:::: is composed of an electric part, ∆''Ψ''<sub>mt</sub>, which is the difference of charge (electrical potential difference) across the inner mitochondrial membrane, and a chemical part, ∆''µ''<sub>H+</sub>/''F'', which stems from the difference of pH (chemical potential difference) across the inner mitochondrial membrane and incorporates the Faraday constant, ''F''. In other words, the protonmotive force is expressed as the sum of two terms, with somewhat complicated symbols in Eq. 1, which can be more easily explained as isomorphic partial protonmotive forces.
:::::: '''Electrical, el:''' ''F<sub>e'',el</sub> = ∆''Ψ''<sub>mt</sub> is the electrical pat of the protonmotive force expressed in units joules per coulomb, i.e., volt [V=J/C], and defined as partial Gibbs energy change per ''motive elementary charge of protons, e'' [C]. ''F<sub>n'',el</sub> = ∆''Ψ''<sub>mt</sub>∙''F'' is the electrical force expressed in units joules per mole [V=J/mol], and defined as partial Gibbs energy change per ''motive amount of charge, n'' [mol].
:::::: '''Chemical, diffusion or dislocation, d:''' ''F<sub>n'',d</sub> = ∆''µ''<sub>H+</sub> is the corresponding chemical part of the protonmotive force expressed in units joules per mole [J/mol], and defined as partial Gibbs energy change per ''motive amount of protons, n'' [mol]. ''F<sub>e'',d</sub> = ∆''µ''<sub>H+</sub>/''F'' is the chemical force expressed in units joules per coulomb, i.e., volt [V=J/C], and defined as partial Gibbs energy change per ''motive amount of protons expressed in units of electric charge, e'' [C].
:::::: '''Faraday constant, ''F'':''' The Faraday constant is the product of the elementary charge and the Avogadro (or Loschmidt) constant, ''F'' = ''e∙NA'' [C/mol], and yields the conversion between protonmotive force, ''F<sub>e</sub>'' = ∆''p''<sub>mt</sub> [J/C], expressed per ''motive charge, e'' [C], and protonmotive force or chemiosmotic potential difference, ''F<sub>n</sub>'' = ∆''p''<sub>mt</sub>∙''F'' [J/mol], expressed per ''motive amount of substance, n'' [mol],
:::::::::::::: ''F<sub>n</sub>'' = ''F<sub>e</sub>'' ∙ ''F'' ; (Eq. 2.1)
:::::::::::::: ''F<sub>e</sub>'' = ''F<sub>e'',el</sub> + ''F<sub>e'',d</sub>  =  ∆''Ψ''<sub>mt</sub> + ∆µH<sup>+</sup>/''F'' ; ''e''-isomorph [J/C=V] (Eq. 2.2)
:::::::::::::: ''F<sub>n</sub>'' = ''F<sub>n'',el</sub> + ''F<sub>n'',d</sub>  =  ∆''Ψ''<sub>mt</sub>∙''F'' + ∆''µ''<sub>H+</sub> ; ''n''-isomorph [J/mol] (Eq. 2.3)
:::: Protonmotive means that protons are moved across the mitochondrial membrane at constant force, and the direction of the translocation is defined in Fig. 2 as H<big>+</big><sub>in</sub> → H<sup>+</sup><sub>out</sub>,
:::::::::::::: ''F<sub>n'',d</sub> = ∆''µ''<sub>H+</sub> = -ln(10)∙''RT''∙∆pH<sub>mt</sub> ; (Eq. 3)
:::: where ''RT'' is the gas constant times absolute temperature. ln(10)∙''RT'' = 5.708 and 5.938 kJ∙mol­<sup>-1</sup> at 25 and 37 °C, respectively. ln(10)∙''RT/F'' = 59.16 and 61.54 mV at 25 and 37 °C, respectively. For a ∆pH of 1 unit, the chemical force (Eq. 3) changes by 6 kJ∙mol­<sup>-1</sup> and the protonmotive force (Eq. 2.2) changes by 0.06 V.
:::: Since ''F'' equals 96.5 (kJ∙mol­<sup>-1</sup>)/V, a membrane potential difference of -0.2 V (Eq. 2.2) equals a chemiosmotic potential difference, ''F<sub>n</sub>'', of 19 kJ∙mol<sup>-1</sup> H<sup>+</sup><sub>out</sub><sub>Subscript text</sub> (Eq. 2.3). Considering a driving force of -470 kJ∙mol<sup>-1</sup> O<sub>2</sub> for oxidation, the thermodynamic limit of the H<sup>+</sup><sub>out</sub>/O<sub>2</sub> ratio is reached at a value of 470/19 = 24, compared to a mechanistic stoichiometry of 20 (H<sup>+</sup><sub>out</sub>/O=10).


:::: ''I thank you in advance for your feedback.''
:::: ''With best wishes,''


Perspective added in Version 17
:::: To provide an overall perspective of mitochondrial physiology we may link cellular bioenergetics to systemic human respiratory activity, without yet addressing cell- and tissue-specific mitochondrial function. A routine O<sub>2</sub> flow of 234 µmol∙s<sup>-1</sup> per individual or flux of 3.3 nmol∙s<sup>-1</sup>∙g<sup>-1</sup> body mass corresponds to -110 W catabolic energy flow at a body mass of 70 kg and -470 kJ/mol O<sub>2</sub>. Considering a cell count of 514∙10<sup>6</sup> cells per g tissue mass and an estimate of 300 mitochondria per cell (Ahluwalia 2017), the average oxygen flow per million cells at ''J<sub>m'',O2peak</sub> of 45 nmol·s<sup>-1</sup>·g<sup>-1</sup> (60 ml O<sub>2</sub>·min<sup>-1</sup>·kg<sup>-1</sup>) is 88 pmol∙s­<sup>-1</sup>∙10­<sup>-6</sup> cells, which compares well with OXPHOS capacity of human fibroblasts (not ETS but the lower OXPHOS capacity is used as a reference; Gnaiger 2014). We can describe our body as the sum of 36∙10<sup>12</sup> cells (36 trillion cells). Mitochondrial fitness of our 11∙10<sup>15</sup> mitochondria (11 quadrillion mt) is indicated if O<sub>2</sub> flow of 0.02 pmol∙s­<sup>-1</sup>∙10<sup>-6</sup> mt­ at rest can be activated to 0.3 pmol∙s<sup>-1</sup>∙10<sup>-6</sup> mt at high ergometric performance.


* 2018-08-16 Erich Gnaiger added Fig. 6 and text in Version 16:
:::: ''Erich Gnaiger''
:::: Fig. 6 summarizes the three coupling states, ETS, LEAK and OXPHOS, and puts them into a schematic context with the corresponding respiratory rates, abbreviated as ''E, L'' and ''P'', respectively. This clarifies that ''E'' may exceed or be equal to ''P'', but ''E'' cannot theoretically be lower than ''P''. ''E<P'' must be discounted as an artefact, which may be caused experimentally by (i) using high and inhibitory uncoupler concentrations (Gnaiger 2008), (ii) non-saturating [ADP] or [Pi] (State 3), (iii) high oligomycin concentrations applied for measurement of ''L'' before titrations of uncoupler, when oligomycin exerts an inhibitory effect on ''E'', or (iv) loss of oxidative capacity during the time course of the respirometric assay with ''E'' measured subsequently to ''P'' (Gnaiger 2014). On the other hand, ''E>P'' is observed in many types of mitochondria and depends on (i) the excess ETS capacity pushing the phosphorylation system (Fig. 1B) to the limit of its capacity of utilizing ∆''p''<sub>mt</sub>, (ii) the pathway control state with single or multiple electron input into the Q-junction and involvement of three or less coupling sites determining the H<sup>+</sup><sub>out</sub>/O<sub>2</sub> coupling stoichiometry (Fig. 2A), and (iii) the biochemical coupling efficiency expressed as (''E-L'')/''E'', since any increase of ''L'' causes an increase of ''P'' upwards to the limit of ''E''. The ''excess E-P'' capacity, ''ExP=E-P'', therefore, provides a sensitive diagnostic indicator of specific injuries of the phosphorylation system, when ''E'' remains constant but ''P'' declines relative to controls (Fig. 6). Substrate cocktails supporting simultaneous convergent electron transfer to the Q-junction for reconstitution of TCA cycle function stimulate ETS capacity, and consequently increase the sensitivity of the ''ExP'' assay.
:::: ''Chair Mitochondrial Physiology Society'' - http://www.mitophysiology.org
 
:::: ''Chair COST Action MitoEAGLE'' - http://www.mitoeagle.org
:::: ... In general, it is inappropriate to use the term ''ATP production'' for the difference of oxygen consumption measured in states ''P'' and ''L''. The difference ''P-L'' is the upper limit of the part of OXPHOS capacity which is ''free'' (corrected for LEAK respiration) and is fully coupled to phosphorylation with a maximum mechanistic stoichoimetry, ''≈P = P-L'' (Fig. 6).
:::: ''Medical University of Innsbruck, Austria''


__TOC__
__TOC__
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::::* Scientific terminology should be general and platform-independent, meeting the demands of all working groups.
::::* Scientific terminology should be general and platform-independent, meeting the demands of all working groups.


=== Abstract ===
:::::::::::: ''‘Every professional group develops its own technical jargon for talking about matters of critical concern. .. People who know a word can share that idea with other members of their group, and a shared vocabulary is part of the glue that holds people together and allows them to create a shared culture’'' (Miller 1991).
:::: Clarity of concepts and consistency of nomenclature is a hallmark of the quality of a research field across its specializations, aimed at facilitating transdisciplinary communication and education. As mitochondrial physiology continues to expand, the necessity for improved harmonization of nomenclature on mitochondrial respiratory states and rates has become apparent. Peter Mitchell’s protonmotive force across the inner mitochondrial membrane, Δ''p''<sub>mt</sub>, establishes the link between electron transfer and phosphorylation of ADP to ATP, and between the electric and chemical components of energy transformation (Δ''Ψ''<sub>mt</sub> and ΔpH). This unifying concept provides the framework for developing a consistent terminology on mitochondrial physiology and bioenergetics. We follow IUPAC guidelines on general terms of physical chemistry, extended by concepts of nonequilibrium thermodynamics and open systems. The nomenclature of respiratory sates in classical bioenergetics (States 1 to 5 in an experimental protocol) is incorporated into a concept-driven constructive terminology to address the meaning of each respiratory state and to focus primarily on the conceptual ‘why’ with clarification of the experimental ‘how’. [[LEAK respiration |LEAK]] states are evaluated to study arrested respiration, when oxygen consumption compensates mainly for the proton leak. [[Oxidative phosphorylation |OXPHOS]] capacity is measured at saturating concentrations of ADP and inorganic phosphate to obtain kinetic reference values for diagnostic applications. The oxidative capacity of the [[electron transfer system]] is determined in the ETS state, revealing the limitation of OXPHOS capacity mediated by the capacity of the phosphorylation system. Development of databases of mitochondrial respiratory control requires the application of strictly defined terms for comparison of respiratory states.


:::::::::::: ''‘Every professional group develops its own technical jargon for talking about matters of critical concern. .. People who know a word can share that idea with other members of their group, and a shared vocabulary is part of the glue that holds people together and allows them to create a shared culture’'' (Miller 1991).
=== Three fundamental coupling states of mitochondrial preparations and residual oxygen consumption ===
::::* '''[[OXPHOS]]'''
::::* '''[[Electron transfer pathway]]'''
::::* '''[[LEAK respiration]]'''
::::* '''[[ROX]]'''


=== Classical terminology for isolated mitochondria ===
=== Classical terminology for isolated mitochondria ===
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::::* '''[[State 3]]'''
::::* '''[[State 3]]'''
::::* '''[[State 4]]'''
::::* '''[[State 4]]'''
=== Three fundamental coupling states of mitochondrial preparations and residual oxygen consumption ===
::::* '''[[OXPHOS]]'''
::::* '''[[ETS]]'''
::::* '''[[LEAK]]'''
::::* '''[[ROX]]'''




=== States and rates ===
=== States and rates ===
::::* '''[[Force |Force, ''F''<sub>tr</sub>]]''': A generalized force (intensive quantity) in thermodynamics or ergodynamics is the partial Gibbs (Helmholtz) energy change per advancement of a transformation (tr).
::::* '''[[Force |Force, ''F''<sub>tr</sub>]]'''
::::* '''[[External flow |External flow, ''I''<sub>ext</sub>]]'''
::::* '''[[External flow |External flow, ''I''<sub>ext</sub>]]'''
::::* '''[[Internal flow |Internal flow, ''I''<sub>int</sub>]]'''
::::* '''[[Internal flow |Internal flow, ''I''<sub>int</sub>]]'''
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::::* '''[[Respiratory state]]'''
::::* '''[[Respiratory state]]'''


[[File:Rate.jpg|right|400px|thumb|Fig. 6. Different meanings of rate may lead to confusion, if the normalization is not sufficiently specified.]]
=== Normalization: fluxes and flows ===
=== Normalization: fluxes and flows ===
::::* Units (important for a database); analogous to electic terms: Flow [C.s<sup>-1</sup>]; Flux [C.s<sup>-1</sup>.m<sup>-2</sup>]; Rate (?)
::::* '''[[Extensive quantity]]'''
::::* '''[[Extensive quantity]]'''
::::* '''[[Oxygen flow |Oxygen flow, ''I''<sub>O2</sub>]]'''
::::* '''[[Oxygen flow |Oxygen flow, ''I''<sub>O2</sub>]]'''
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::::* '''[[Specific quantity]]'''
::::* '''[[Specific quantity]]'''
::::* '''[[Intensive quantity]]'''
::::* '''[[Intensive quantity]]'''


=== List of selected terms and symbols ===
=== List of selected terms and symbols ===
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::::* '''[[Isolated mitochondria |Isolated mitochondria, imt]]'''
::::* '''[[Isolated mitochondria |Isolated mitochondria, imt]]'''
::::* '''Mitochondria, mt''' (Greek mitos: thread; chondros: granule) are small structures within cells, which function in cell respiration as powerhouses or batteries. Mitochondria belong to the bioblasts of Richard Altmann (1894). Abbreviation: mt, as generally used in mtDNA. Singular: mitochondrion (bioblast); plural: mitochondria (bioblasts).
::::* '''Mitochondria, mt''' (Greek mitos: thread; chondros: granule) are small structures within cells, which function in cell respiration as powerhouses or batteries. Mitochondria belong to the bioblasts of Richard Altmann (1894). Abbreviation: mt, as generally used in mtDNA. Singular: mitochondrion (bioblast); plural: mitochondria (bioblasts).
::::* '''[[Mitochondrial inner membrane |Mitochondrial inner membrane, mt-im]]'''
::::* '''[[Mitochondrial inner membrane]]'''
::::* '''[[Mitochondrial outer membrane |Mitochondrial outer membrane, mt-om]]'''
::::* '''[[Mitochondrial outer membrane]]'''
::::* '''[[Mitochondrial matrix |Mitochondrial matrix, mt-matrix]]'''
::::* '''[[Mitochondrial matrix]]'''
::::* '''[[Mitochondrial membrane potential |Mitochondrial membrane potential, mtMP, Δ''ψ''<sub>mt</sub>]]
::::* '''[[Mitochondrial membrane potential]]
::::* '''Mitochondrial preparations, mtprep''' are isolated mitochondria (imt), tissue homogenate (thom), mechanically or chemically permeabilized tissue (permeabilized fibres, pfi) or permeabilized cells (pce). In these preparations the cell membranes are either removed (imt and smtp) or mechanically (thom) and chemically permeabilized (pfi), while the mitochondrial functional integrity and to a large extent the mt-structure is maintained.
::::* '''Mitochondrial preparations, mtprep''' are isolated mitochondria (imt), tissue homogenate (thom), mechanically or chemically permeabilized tissue (permeabilized fibres, pfi) or permeabilized cells (pce). In these preparations the cell membranes are either removed (imt and smtp) or mechanically (thom) and chemically permeabilized (pfi), while the mitochondrial functional integrity and to a large extent the mt-structure is maintained.
::::* '''[[Mitochondrial respiration]]'''
::::* '''[[Mitochondrial respiration]]'''
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::::* '''[[Proton pump]]'''
::::* '''[[Proton pump]]'''
::::* '''[[Proton slip]]'''
::::* '''[[Proton slip]]'''
::::* '''Protonmotive force, Δ''p''<sub>mt</sub>'''
::::* '''Protonmotive force'''
::::* '''[[Tissue homogenate |Tissue homogenate, thom]]'''
::::* '''[[Tissue homogenate |Tissue homogenate, thom]]'''
::::* '''[[Uncoupler |Uncoupler, U]]'''
::::* '''[[Uncoupler |Uncoupler, U]]'''




== Mitochondrial respiratory control: pathway states in mt-preparations ==
== Mitochondrial pathways and respiratory control in mt-preparations ==
::: '''MitoEAGLE recommendations Part 2.'''
::: '''Building blocks of mitochondrial physiology Part 2.'''


:::::::::::: ''‘It is essential to define both the substrate and ADP levels in order to identify the steady-state condition of the mitochondria during the experiment’'' (Chance and Williams, 1956).
:::::::::::: ''‘It is essential to define both the substrate and ADP levels in order to identify the steady-state condition of the mitochondria during the experiment’'' (Chance and Williams, 1956).
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::::* The mitochondrial respiratory system
::::* The mitochondrial respiratory system
::::* Substrates and inhibitors
::::* Substrates and inhibitors
::::* Switch to pathway-related nomenclature instead of enzyme-linked terminology (N/NS/S versus CI/CI+II/CII)
::::* Switch to pathway-related nomenclature instead of enzyme-linked terminology (N/NS/S versus CI/CI&II/CII)




== Mitochondrial respiratory control: cell respiration ==
== Mitochondrial respiratory control: cell respiration ==
::: '''MitoEAGLE recommendations Part 3.'''
::: '''Building blocks of mitochondrial physiology Part 3.'''


::: '''Intact cells versus mitochondrial preparations'''
::: '''Living cells versus mitochondrial preparations'''
::::* '''[[Intact cells |Intact cells, ce]]'''
::::* '''[[Living cells |Living cells, ce]]'''
::::* '''[[Basal respiration]]'''
::::* '''[[Basal respiration]]'''
::::* '''[[Cell respiration]]'''
::::* '''[[Cell respiration]]'''
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::::* '''[[ROUTINE respiration |ROUTINE state, state ''R'']]''': ROUTINE respiration of intact, viable cells is regulated according to physiological activity, at intracellular non-saturating ADP levels. ''R'' increases under various conditions of activation. When incubated in culture medium, cells maintain a ROUTINE level of activity, ''R'' (ROUTINE mitochondrial respiration; corrected for residual oxygen consumption due to oxidative side reactions). ROUTINE activity may include aerobic energy requirements for cell growth and is thus fundamentally different from the definition of basal metabolic rate (BMR). When incubated for short experimental periods in a medium devoid of fuel substrates, the cells respire solely on endogenous substrates at the corresponding state of ROUTINE activity, e<sub>''R''</sub> (e, endogenous substrate supply).
::::* '''[[ROUTINE respiration |ROUTINE state, state ''R'']]''': ROUTINE respiration of intact, viable cells is regulated according to physiological activity, at intracellular non-saturating ADP levels. ''R'' increases under various conditions of activation. When incubated in culture medium, cells maintain a ROUTINE level of activity, ''R'' (ROUTINE mitochondrial respiration; corrected for residual oxygen consumption due to oxidative side reactions). ROUTINE activity may include aerobic energy requirements for cell growth and is thus fundamentally different from the definition of basal metabolic rate (BMR). When incubated for short experimental periods in a medium devoid of fuel substrates, the cells respire solely on endogenous substrates at the corresponding state of ROUTINE activity, e<sub>''R''</sub> (e, endogenous substrate supply).


:::: It is difficult to stimulate living cells to maximum OXPHOS activity, since ADP and inorganic phosphate do not equilibrate across intact plasma membranes, and thus saturating concentrations of these metabolites can hardly be achieved in living cells. LEAK and ETS states, however, can be induced in viable cells with application of inhibitors of the phosphorylation system and uncouplers, respectively, due to the fact that cell membranes are highy permeable for these substances. External fuel substrates are taken up by living cells to various extents, and intracellular metabolism of exogenous and endogenous substrates supports mitochondrial respiration with a physiological substrate supply. In contrast, mt-preparations depend on the external supply of fuel substrates which support the electron transfer system with reducing equivalents. ETS competence of external substrates is required for all coupling states of mt-preparations (''L, P, E'') and depends on (i) transport of substrates across the inner mt-membrane or oxidation by dehydrogenases located on the outer face of the inner mt-membrane (e.g. glycerophosphate dehydrogenase complex, CGpDH), (ii) oxidation in the mt-matrix (TCA cycle dehydrogenases and other matrix dehydrogenases, e.g. mtGDH) or on the inner face of the inner mt-membrane (succinate dehydrogenase, CII), (iii) oxidation of substrates without accumulation of inhibitory endproducts (e.g. oxaloacetate inhibiting succinate dehydrogenase; NADH and oxaloacetate inhibiting malate dehydrogenase), and (iv) electron transfer through the membrane-bound ETS (mETS). Endproducts must be either easily exported from the matrix across the inner mt-membrane (e.g. malate formed from succinate via fumarate), or metabolized in the TCA cycle (e.g. malate-derived oxaloacetate forming citrate in the presence of external pyruvate&malate).
:::: It is difficult to stimulate living cells to maximum OXPHOS activity, since ADP and inorganic phosphate do not equilibrate across intact plasma membranes, and thus saturating concentrations of these metabolites can hardly be achieved in living cells. LEAK and ET-pathway states, however, can be induced in viable cells with application of inhibitors of the phosphorylation system and uncouplers, respectively, due to the fact that cell membranes are highy permeable for these substances. External fuel substrates are taken up by living cells to various extents, and intracellular metabolism of exogenous and endogenous substrates supports mitochondrial respiration with a physiological substrate supply. In contrast, mt-preparations depend on the external supply of fuel substrates which support the electron transfer-pathway with reducing equivalents. ET-pathway competence of external substrates is required for all coupling states of mt-preparations (''L, P, E'') and depends on (i) transport of substrates across the inner mt-membrane or oxidation by dehydrogenases located on the outer face of the inner mt-membrane (e.g. glycerophosphate dehydrogenase complex, CGpDH), (ii) oxidation in the mt-matrix (TCA cycle dehydrogenases and other matrix dehydrogenases, e.g. mtGDH) or on the inner face of the inner mt-membrane (succinate dehydrogenase, CII), (iii) oxidation of substrates without accumulation of inhibitory endproducts (e.g. oxaloacetate inhibiting succinate dehydrogenase; NADH and oxaloacetate inhibiting malate dehydrogenase), and (iv) electron transfer through the membrane-bound ET-pathway (mETS). Endproducts must be either easily exported from the matrix across the inner mt-membrane (e.g. malate formed from succinate via fumarate), or metabolized in the TCA cycle (e.g. malate-derived oxaloacetate forming citrate in the presence of external pyruvate&malate).




== Mitochondrial respiratory control: coupling control ratios and control factors ==
== Mitochondrial respiratory control: coupling control ratios and control factors ==
::: '''MitoEAGLE recommendations Part 4.'''
::: '''Building blocks of mitochondrial physiology Part 4.'''


::::* '''[[Coupling control factor |Coupling control factor, ''CCF'']]'''
::::* '''[[Coupling control factor |Coupling control factor, ''CCF'']]'''
::::* '''[[Coupling control ratio |Coupling control ratio, ''CCR'']]'''
::::* '''[[Coupling-control ratio |Coupling-control ratio, ''CCR'']]'''


::::* '''[[ETS coupling efficiency |ETS coupling efficiency, ''j<sub>≈E</sub>'']]''': ''j<sub>≈E</sub>'' = ''≈E/E'' = (''E-L'')/''E'' = 1-''L/E''
::::* '''[[E-L coupling efficiency|E-L coupling efficiency, ''j<sub>≈E</sub>'']]''': ''j<sub>≈E</sub>'' = ''≈E/E'' = (''E-L'')/''E'' = 1-''L/E''
::::* '''[[LEAK control ratio |LEAK control ratio, ''L/E'']]'''
::::* '''[[LEAK-control ratio |LEAK-control ratio, ''L/E'']]'''


::::* '''[[Excess E-P capacity factor |Excess ''E-P'' capacity factor, ''j<sub>ExP</sub>'']]''': ''j<sub>Exp</sub>'' = (''E-P'')/''E'' = 1-''P/E''
::::* '''[[Excess E-P capacity factor |Excess ''E-P'' capacity factor, ''j<sub>ExP</sub>'']]''': ''j<sub>Exp</sub>'' = (''E-P'')/''E'' = 1-''P/E''
::::* '''[[OXPHOS control ratio |OXPHOS control ratio, ''P/E'']]'''
::::* '''[[OXPHOS-control ratio |OXPHOS-control ratio, ''P/E'']]'''


::::* '''[[OXPHOS coupling efficiency |OXPHOS coupling efficiency, ''j<sub>≈P</sub>'']]''': ''j<sub>≈P</sub>'' = ''≈P/P'' = (''P-L'')/''P'' = 1-''L/P''
::::* '''[[OXPHOS-coupling efficiency |OXPHOS-coupling efficiency, ''j<sub>≈P</sub>'']]''': ''j<sub>≈P</sub>'' = ''≈P/P'' = (''P-L'')/''P'' = 1-''L/P''
::::* '''[[L/P coupling control ratio |''L/P'' coupling control ratio, ''L/P'']]'''
::::* '''[[L/P coupling control ratio |''L/P'' coupling control ratio, ''L/P'']]'''
::::* '''[[Respiratory acceptor control ratio |Respiratory acceptor control ratio, RCR]]''': RCR = State 4/State 3
::::* '''[[Respiratory acceptor control ratio |Respiratory acceptor control ratio, RCR]]''': RCR = State 4/State 3
Line 193: Line 131:
::::* '''[[NetOXPHOS control ratio |netOXPHOS control ratio, ''≈P/E'']]''': ''≈P/E'' = (''P-L'')/''E''
::::* '''[[NetOXPHOS control ratio |netOXPHOS control ratio, ''≈P/E'']]''': ''≈P/E'' = (''P-L'')/''E''


::::* '''[[Uncoupling control ratio |Uncoupling control ratio, UCR]]''': UCR = ''E/R''
::::* '''[[Uncoupling-control ratio |Uncoupling-control ratio, UCR]]''': UCR = ''E/R''




== Action ==
== Action ==
::::» '''WG1 Action''' - [[WG1 MITOEAGLE protocols, terminology, documentation]]: Standard operating procedures and user requirement document: Protocols, terminology, documentation
::::» '''WG1 Action''' - [[WG1 MitoEAGLE protocols, terminology, documentation]]: Standard operating procedures and user requirement document: Protocols, terminology, documentation
::::» [[MITOEAGLE protocols, terminology, documentation |WG1 Project application]]
::::» [[MitoEAGLE protocols, terminology, documentation |WG1 Project application]]
::::» Gnaiger E, Aasander Frostner E, Abdul Karim N, Abumrad NA, Acuna-Castroviejo D, Adiele RC et al (2019) Mitochondrial respiratory states and rates. MitoFit Preprint Arch doi:10.26124/mitofit:190001. - [[Gnaiger 2019 MitoFit Preprints |»Bioblast link«]]


 
::::» [[MitoEAGLE: Respiratory states |Pre-publication: Mitochondrial respiratory control states]]
::::» [[MITOEAGLE: Respiratory states |Pre-publication: Mitochondrial respiratory control states]]
::::» [[MitoPedia: Respiratory control ratios]]
::::» [[MitoPedia: Respiratory control ratios]]
::::» [[MitoPedia: SUIT]]
::::» [[MitoPedia: SUIT]]
::::» 2017-07 [[MiPschool Obergurgl 2017]]
::::» 2017-03 [[MITOEAGLE Barcelona 2017]]
::::» 2016-11 [[MITOEAGLE 2016 Verona IT]]

Latest revision as of 23:40, 25 January 2021


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COST Action CA15203 (2016-2021): MitoEAGLE
Evolution-Age-Gender-Lifestyle-Environment: mitochondrial fitness mapping


The protonmotive force and respiratory control


Building blocks of mitochondrial physiology MitoEAGLE recommendations Part 1.



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» Manuscript phases and versions

Manuscript phases and versions - an open-access apporach

COST Action MitoEAGLE
This manuscript on ‘Mitochondrial respiratory states and rates’ is a position statement in the frame of COST Action CA15203 MitoEAGLE. The list of coauthors evolved beyond phase 1 in the bottom-up spirit of COST.
The global MitoEAGLE network made it possible to collaborate with a large number of coauthors to reach consensus on the present manuscript. Nevertheless, we do not consider scientific progress to be supported by ‘declaration’ statements (other than on ethical or political issues). Our manuscript aims at providing arguments for further debate rather than pushing opinions. We hope to initiate a much broader process of discussion and want to raise the awareness on the importance of a consistent terminology for reporting of scientific data in the field of bioenergetics, mitochondrial physiology and pathology. Quality of research requires quality of communication. Some established researchers in the field may not want to re-consider the use of jargon which has become established despite deficiencies of accuracy and meaning. In the long run, superior standards will become accepted. We hope to contribute to this evolutionary process, with an emphasis on harmonization rather than standardization.
  • Phase 1: The protonmotive force and respiratory control
» The protonmotive force and respiratory control - Discussion
» MitoEAGLE preprint 2017-09-21 - Discussion
  • Phase 2: Mitochondrial respiratory states and rates: Building blocks of mitochondrial physiology Part 1
» MitoEAGLE Task Group States and rates - Discussion
  • Phase 4: Journal submission
  • Target: CELL METABOLISM, aiming at indexing by The Web of Science and PubMed.
Coauthors
  • 2017-09-21 Version 01: 105 coauthors
  • 2017-10-15 Version 10: 131 coauthors
  • 2018-01-18 Version 20: 168 coauthors
  • 2018-02-26 Version 30: 225 coauthors
  • 2018-08-20 Version 40: 350 coauthors - EBEC Poster
  • 2018-10-17 Version 44: 426 coauthors - MiPschool Tromso-Bergen 2018
  • 2018-12-12 Version 50: 517 coauthors - Submission to the preprint server bioRxiv not successful
  • 2019-02-12 Preprint version 1: 530 coauthors
  • 2019-03-15 Preprint version 2: 533 coauthors
  • 2019-04-24 Preprint version 3: 533 coauthors
  • 2019-05-20 Preprint version 4: 542 coauthors
  • 2019-07-24 Preprint version 5: 612 coauthors
  • 2019-08-30 Preprint version 6: 622 coauthors - Preprint publication doi:10.26124/mitofit:190001.v6
  • BEC 2020.1. - Gnaiger Erich et al ― MitoEAGLE Task Group (2020) Mitochondrial physiology. Bioenerg Commun 2020.1. doi:10.26124/bec:2020-0001.v1. - »Bioblast link«



Open invitation

This manuscript on ‘The protonmotive force and respiratory control’ is a position statement in the frame of COST Action CA15203 MitoEAGLE. The list of coauthors evolved from MitoEAGLE Working Group Meetings and a bottom-up spirit of COST: This is an open invitation to scientists and students to join as coauthors, to provide a balanced view on mitochondrial respiratory control, a fundamental introductory presentation of the concept of the protonmotive force, and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. We plan a series of follow-up reports by the expanding MitoEAGLE Network, to increase the scope of consensus-oriented recommendations and facilitate global communication and collaboration.
  • We continue to invite comments and suggestions on the MitoEAGLE preprint (phase 2; until October 12), particularly if you are an early career investigator adding an open future-oriented perspective, or an established scientist providing a balanced historical basis. Your critical input into the quality of the manuscript will be most welcome, improving our aims to be educational, general, consensus-oriented, and practically helpful for students working in mitochondrial respiratory physiology.
  • Please feel free to focus on a particular section in terms of direct input and references, while evaluating the entire scope of the manuscript from the perspective of your expertise.
  • Your comments will be largely posted on the discussion page of the MitoEAGLE preprint website. If you prefer to submit comments in the format of a referee's evaluation rather than a contribution as a coauthor, I will be glad to distribute your views to the updated list of coauthors for a balanced response. We would ask for your consent on this open bottom-up procedure.
We organize a MitoEAGLE session linked to our series of communications at the MiPconference Nov 2017 in Hradec Kralove in close association with the MiPsociety (where you hopefully will attend) and at EBEC 2018 in Budapest.
» MiP2017_Hradec_Kralove_CZ
I thank you in advance for your feedback.
With best wishes,


Erich Gnaiger
Chair Mitochondrial Physiology Society - http://www.mitophysiology.org
Chair COST Action MitoEAGLE - http://www.mitoeagle.org
Medical University of Innsbruck, Austria
Scope
  • Target a broad audience – introduce the new generation of investigators.
  • List of terms including historical terms; abbreviations (mtDNA, mt to abbreviate mitochondr*); OXPHOS capacity versus State 3 (discuss saturating ADP/Pi .. concentrations).
  • Protonmotive force: not a force defined in physics, but an isomorphic force of statistical and nonequilibrium thermodynamics.
  • Flux and flow: clarification of normalization.
  • Scientific terminology should be general and platform-independent, meeting the demands of all working groups.
‘Every professional group develops its own technical jargon for talking about matters of critical concern. .. People who know a word can share that idea with other members of their group, and a shared vocabulary is part of the glue that holds people together and allows them to create a shared culture’ (Miller 1991).

Three fundamental coupling states of mitochondrial preparations and residual oxygen consumption

Classical terminology for isolated mitochondria

‘When a code is familiar enough, it ceases appearing like a code; one forgets that there is a decoding mechanism. The message is identical with its meaning’ (Hofstadter 1979).


States and rates

Normalization: fluxes and flows

List of selected terms and symbols


Mitochondrial pathways and respiratory control in mt-preparations

Building blocks of mitochondrial physiology Part 2.
‘It is essential to define both the substrate and ADP levels in order to identify the steady-state condition of the mitochondria during the experiment’ (Chance and Williams, 1956).
  • The mitochondrial respiratory system
  • Substrates and inhibitors
  • Switch to pathway-related nomenclature instead of enzyme-linked terminology (N/NS/S versus CI/CI&II/CII)


Mitochondrial respiratory control: cell respiration

Building blocks of mitochondrial physiology Part 3.
Living cells versus mitochondrial preparations
  • Living cells, ce
  • Basal respiration
  • Cell respiration
  • Resting metabolic rate
  • ROUTINE state, state R: ROUTINE respiration of intact, viable cells is regulated according to physiological activity, at intracellular non-saturating ADP levels. R increases under various conditions of activation. When incubated in culture medium, cells maintain a ROUTINE level of activity, R (ROUTINE mitochondrial respiration; corrected for residual oxygen consumption due to oxidative side reactions). ROUTINE activity may include aerobic energy requirements for cell growth and is thus fundamentally different from the definition of basal metabolic rate (BMR). When incubated for short experimental periods in a medium devoid of fuel substrates, the cells respire solely on endogenous substrates at the corresponding state of ROUTINE activity, eR (e, endogenous substrate supply).
It is difficult to stimulate living cells to maximum OXPHOS activity, since ADP and inorganic phosphate do not equilibrate across intact plasma membranes, and thus saturating concentrations of these metabolites can hardly be achieved in living cells. LEAK and ET-pathway states, however, can be induced in viable cells with application of inhibitors of the phosphorylation system and uncouplers, respectively, due to the fact that cell membranes are highy permeable for these substances. External fuel substrates are taken up by living cells to various extents, and intracellular metabolism of exogenous and endogenous substrates supports mitochondrial respiration with a physiological substrate supply. In contrast, mt-preparations depend on the external supply of fuel substrates which support the electron transfer-pathway with reducing equivalents. ET-pathway competence of external substrates is required for all coupling states of mt-preparations (L, P, E) and depends on (i) transport of substrates across the inner mt-membrane or oxidation by dehydrogenases located on the outer face of the inner mt-membrane (e.g. glycerophosphate dehydrogenase complex, CGpDH), (ii) oxidation in the mt-matrix (TCA cycle dehydrogenases and other matrix dehydrogenases, e.g. mtGDH) or on the inner face of the inner mt-membrane (succinate dehydrogenase, CII), (iii) oxidation of substrates without accumulation of inhibitory endproducts (e.g. oxaloacetate inhibiting succinate dehydrogenase; NADH and oxaloacetate inhibiting malate dehydrogenase), and (iv) electron transfer through the membrane-bound ET-pathway (mETS). Endproducts must be either easily exported from the matrix across the inner mt-membrane (e.g. malate formed from succinate via fumarate), or metabolized in the TCA cycle (e.g. malate-derived oxaloacetate forming citrate in the presence of external pyruvate&malate).


Mitochondrial respiratory control: coupling control ratios and control factors

Building blocks of mitochondrial physiology Part 4.


Action

» WG1 Action - WG1 MitoEAGLE protocols, terminology, documentation: Standard operating procedures and user requirement document: Protocols, terminology, documentation
» WG1 Project application
» Gnaiger E, Aasander Frostner E, Abdul Karim N, Abumrad NA, Acuna-Castroviejo D, Adiele RC et al (2019) Mitochondrial respiratory states and rates. MitoFit Preprint Arch doi:10.26124/mitofit:190001. - »Bioblast link«
» Pre-publication: Mitochondrial respiratory control states
» MitoPedia: Respiratory control ratios
» MitoPedia: SUIT
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