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Complex II ambiguities

From Bioblast

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Complex II ambiguities


Two ambiguities or misconceptions around respiratory Complex II (CII) have their roots in the narrative that reduced coenzymes (NADH and FADH2) feed electrons from the tricarboxylic acid (TCA) cycle into the mitochondrial electron transfer system. In graphical representations propagating the first ambiguity, succinate dehydrogenase or CII in the canonical (forward) TCA cycle is shown to reduce FAD to FADH2 (correct), yet CII in the membrane-bound electron transfer system (ETS) is paradoxically represented as the site of oxidation of FADH2 to FAD. With minor expansion of the tale on electron transfer from FADH2 into CII, we arrive at the misconception that FADH2 generated by electron transferring flavoprotein (CETF) in fatty acid oxidation and by mitochondrial glycerophosphate dehydrogenase (CGpDH) feeds electrons into the ETS through CII. For clarification, recall that NADH and succinate formed in the TCA cycle in the mitochondrial matrix are the upstream substrates of Complexes CI and CII, whereas the reduced flavin groups FMNH2 of flavin mononucleotide and FADH2 of flavin adenine dinucleotide are products of CI and CII, respectively, with downstream electron flow from CI and CII into the Q-junction. CETF and CGpDH feed electrons into the Q-junction convergent with but not through CII into the Q-junction. Numerous Complex II ambiguities in the literature with discrepancies in graphical representations and text require quality control to secure scientific standards in current communications on bioenergetics.

Abbreviation: CII ambiguities

Communicated by Gnaiger E (2023-03-03) last update 2023-03-21

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Arnold, Finley 2022 Fig1.png
Ref. [1] Arnold PK, Finley LWS (2023) Regulation and function of the mammalian tricarboxylic acid cycle. J Biol Chem 299:102838. - »Bioblast link«

FADH2 and FMNH2 in the S- and N-pathways

Respiratory Complex CII participates both in the membrane-bound electron transfer system (membrane-ETS) and TCA cycle (matrix-ETS plus CII; Gnaiger et al 2020). Branches of electron transfer from the reduced coenzyme NADH of nicotinamide adenine dinucleotide N and succinate S converge at the Q-junction in the ETS (Figure ;a modified from Gnaiger 2020).
The reduced flavin groups FADH2 of flavin adenine dinucleotide and FMNH2 of flavin mononucleotide are at functionally comparable levels in the electron transfer to Q from CII and CI, respectively, just as succinate and NADH are the comparable reduced substrates of CII and CI, respectively (Gnaiger 2020). In CII the oxidized form FAD is reduced by succinate to the product FADH2 and the oxidized product fumarate in the TCA cycle. In CI FMN is reduced by NADH forming FMNH2 and the oxidized NAD+. FADH2 and FMNH2 are reoxidized downstream in CII and CI by electron transfer to Q in the membrane-bound ETS (Figure b).
Ref. [2] Gnaiger E (2020) Mitochondrial pathways and respiratory control. An introduction to OXPHOS analysis. 5th ed.
Ref. [3] Gnaiger E et al ― MitoEAGLE Task Group (2020) Mitochondrial physiology.

The source and consequence of Complex II ambiguities

Ambiguities emerge if the presentation of a concept is vague to an extent that allows for equivocal interpretations. As a consequence of ambiguous representations, even a basically clear and quite simple concept may be communicated further without appropriate reflection as an erroneous divergence from an established truth. The following quotes from Cooper (2000) provide an example.
Cooper 2000 Sunderland 10-9.png
Ref. [4] Cooper GM (2000) The cell: a molecular approach. 2nd edition. Sunderland (MA): Sinauer Associates Available from: - »Bioblast link«
(1) 'Electrons from NADH enter the electron transport chain in complex I, .. A distinct protein complex (complex II), which consists of four polypeptides, receives electrons from the citric acid cycle intermediate, succinate (Figure 10.9). These electrons are transferred to FADH2, rather than to NADH, and then to coenzyme Q.'
Comment: Here, the frequent comparison is made between FADH2 (linked to CII) and NADH (linked to CI).
(2) 'In contrast to the transfer of electrons from NADH to coenzyme Q at complex I, the transfer of electrons from FADH2 to coenzyme Q is not associated with a significant decrease in free energy and, therefore, is not coupled to ATP synthesis.'
Comment: Note that CI is in the path of the transfer of electrons from NADH to coenzyme Q. In contrast, the transfer of electrons from FADH2 to coenzyme Q is downstream of CII. Thus even a large Gibbs force ('decrease in free energy') in FADH2→Q would fail to drive the coupled process of proton translocation through CII, since the Gibbs force in S→FADH2 is missing. (In parentheses: None of these steps are coupled to ATP synthesis. Redox-driven proton translocation should not be confused with pmF-driven phosphorylation of ADP).
(3) 'Electrons from succinate enter the electron transport chain via FADH2 in complex II. They are then transferred to coenzyme Q and carried through the rest of the electron transport chain ..'
Comment: The ambiguity is caused by a lack of unequivocal definition of the electron transfer system ('electron transport chain'). CII receives electrons (1) from succinate, yet it is suggested that electrons (from succinate) enter the electron transport chain (3) via FADH2 in complex II. Then two contrasting definitions are implied of the term 'electron transport chain' or better membrane-bound electron transfer system, membrane-ETS. (a) If CII is part of the membrane-ETS, then electrons enter the membrane-ETS from succinate (1) but not from FADH2. (b) If electrons enter the 'electron transport chain' via FADH2 in Complex II (3), then CII would be upstream and hence not part of the membrane-ETS (to which conclusion, obviously - see Figure - nobody would agree). Dismissing concept (b) of the membrane-ETS, then remains the ambiguity, if electrons enter the membrane-ETS from FADH2 (3, wrong) or from succinate (1, correct).

FADH2 - FAD confusion in the S-pathway

FADH2 appears in several publications as the substrate of CII in the electron transfer system linked to succinate oxidation. It is surprising that this error is widely propagated particularly in the most recent literature. For clarification, see Gnaiger (2020) page 48.
The following examples are listed chronologically and illustrate
(1) ambiguities in graphical representations: FADH2 is the product and substrate of CII in the same figure, e.g. DeBerardinis, Chandel (2016);
(2) evolving errors in graphical representations: e.g. from Figure 6 (ambiguity) to Figure 1 (error) in Chandel (2021);
(3) ambiguities with discrepancies between graphical representation and text: e.g. Figure 1 (error) and text in Fisher-Wellman, Neufer (2012) - 'Reducing equivalents (NADH, FADH2) provide electrons that flow through complex I, the ubiquinone cycle (Q/QH2), complex III, cytochrome c, complex IV, and to the final acceptor O2 to form water' (correct);
(4) simple graphical errors: e.g. Brownlee (2001), Yépez et al (2018), Chen et al (2022); to
(5) propagation of the error in the graphical representation solidified by text: e.g. Arnold, Finley (2022) with the following quotes:
  • 'SDH reduces FAD to FADH2, which donates its electrons to complex II';
  • 'each complete turn of the TCA cycle generates three NADH and one FADH2 molecules, which donate their electrons to complex I and complex II, respectively';
  • 'complex I and complex II oxidize NADH and FADH2, respectively'.
Arnold, Finley 2022 CORRECTION.png
Ref. [1] Arnold PK, Finley LWS (2023) Regulation and function of the mammalian tricarboxylic acid cycle. J Biol Chem 299:102838. - »Bioblast link«

Chen 2022 Am J Physiol Cell Physiol CORRECTION.png
Ref. [5] Chen CL, Zhang L, Jin Z, Kasumov T, Chen YR (2022) Mitochondrial redox regulation and myocardial ischemia-reperfusion injury. Am J Physiol Cell Physiol 322:C12-23. - »Bioblast link«

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Ref. [6] Turton N, Cufflin N, Dewsbury M, Fitzpatrick O, Islam R, Watler LL, McPartland C, Whitelaw S, Connor C, Morris C, Fang J, Gartland O, Holt L, Hargreaves IP (2022) The biochemical assessment of mitochondrial respiratory chain disorders. Int J Mol Sci 23:7487. - »Bioblast link«

Ahmad 2022 StatPearls CORRECTION.png
Ref. [7] Ahmad M, Wolberg A, Kahwaji CI (2022) Biochemistry, electron transport chain. StatPearls Publishing StatPearls [Internet]. Treasure Island (FL) - »Bioblast link«

Yuan 2022 Oxid Med Cell Longev CORRECTION.png
Ref. [8] Yuan Q, Zeng ZL, Yang S, Li A, Zu X, Liu J (2022) Mitochondrial stress in metabolic inflammation: modest benefits and full losses. Oxid Med Cell Longev 2022:8803404. - »Bioblast link«

Chandel 2021 Cold Spring Harb Perspect Biol CORRECTION.png
Ref. [9] Chandel NS (2021) Mitochondria. Cold Spring Harb Perspect Biol 13:a040543. - »Bioblast link«

Ref. [10] Yin M, O'Neill LAJ (2021) The role of the electron transport chain in immunity. FASEB J 35:e21974. - »Bioblast link«
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Ref. [11] Missaglia S, Tavian D, Angelini C (2021) ETF dehydrogenase advances in molecular genetics and impact on treatment. Crit Rev Biochem Mol Biol 56:360-72. - »Bioblast link«

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Ref. [12] Gasmi A, Peana M, Arshad M, Butnariu M, Menzel A, Bjørklund G (2021) Krebs cycle: activators, inhibitors and their roles in the modulation of carcinogenesis. Arch Toxicol 95:1161-78. - »Bioblast link«

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Ref. [13] Turton N, Bowers N, Khajeh S, Hargreaves IP, Heaton RA (2021) Coenzyme Q10 and the exclusive club of diseases that show a limited response to treatment. Expert Opinion on Orphan Drugs 9:151-60. - »Bioblast link«

Martinez-Reyes, Chandel 2020 CORRECTION.png
Ref. [14] Martínez-Reyes I, Chandel NS (2020) Mitochondrial TCA cycle metabolites control physiology and disease. Nat Commun 11:102. - »Bioblast link«

Raimondi 2020 Br J Cancer CORRECTION.png
Ref. [15] Raimondi V, Ciccarese F, Ciminale V (2020) Oncogenic pathways and the electron transport chain: a dangeROS liaison. Br J Cancer 122:168-81. - »Bioblast link«

Morelli 2019 Open Biol CORRECTION.png
Ref. [16] Morelli AM, Ravera S, Calzia D, Panfoli I (2019) An update of the chemiosmotic theory as suggested by possible proton currents inside the coupling membrane. Open Biol 9:180221. - »Bioblast link«

Lewis 2019 CORRECTION.png
Ref. [17] Lewis MT, Kasper JD, Bazil JN, Frisbee JC, Wiseman RW (2019) Quantification of mitochondrial oxidative phosphorylation in metabolic disease: application to Type 2 diabetes. Int J Mol Sci 20:5271. - »Bioblast link«

Sarmah 2019 Transl Stroke Res CORRECTION.png
Ref. [18] Sarmah D, Kaur H, Saraf J, Vats K, Pravalika K, Wanve M, Kalia K, Borah A, Kumar A, Wang X, Yavagal DR, Dave KR, Bhattacharya P (2019) Mitochondrial dysfunction in stroke: implications of stem cell therapy. Transl Stroke Res doi: 10.1007/s12975-018-0642-y - »Bioblast link«
Yepez 2018 PLOS One Fig1B.jpg
Ref. [19] Yépez VA, Kremer LS, Iuso A, Gusic M, Kopajtich R, Koňaříková E, Nadel A, Wachutka L, Prokisch H, Gagneur J (2018) OCR-Stats: Robust estimation and statistical testing of mitochondrial respiration activities using Seahorse XF Analyzer. PLOS ONE 13:e0199938. - »Bioblast link«

Chowdhury 2018 Oxid Med Cell Longev CORRECTION.png
Ref. [20] Roy Chowdhury S, Banerji V (2018) Targeting mitochondrial bioenergetics as a therapeutic strategy for chronic lymphocytic leukemia. Oxid Med Cell Longev 2018:2426712. - »Bioblast link«

De Villiers 2018 Adv Exp Med Biol CORRECTION.png
Ref. [21] de Villiers D, Potgieter M, Ambele MA, Adam L, Durandt C, Pepper MS (2018) The role of reactive oxygen species in adipogenic differentiation. Adv Exp Med Biol 1083:125-144. - »Bioblast link«

Zhang 2018 Mil Med Res CORRECTION.png
Ref. [22] Zhang H, Feng YW, Yao YM (2018) Potential therapy strategy: targeting mitochondrial dysfunction in sepsis. Mil Med Res 5:41. - »Bioblast link«

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Ref. [23] Polyzos AA, McMurray CT (2017) The chicken or the egg: mitochondrial dysfunction as a cause or consequence of toxicity in Huntington's disease. Mech Ageing Dev 161:181-97. - »Bioblast link«

Jones, Bennett 2017 Chapter 4 CORRECTION.png
Ref. [24] Jones PM, Bennett MJ (2017) Chapter 4 - Disorders of mitochondrial fatty acid β-oxidation. Elsevier In: Garg U, Smith LD , eds. Biomarkers in inborn errors of metabolism. Clinical aspects and laboratory determination:87-101. - »Bioblast link«

DeBerardinis, Chandel 2016 CORRECTION.png
Ref. [25] DeBerardinis RJ, Chandel NS (2016) Fundamentals of cancer metabolism. Sci Adv 2:e1600200. - »Bioblast link«

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Ref. [26] Nsiah-Sefaa A, McKenzie M (2016) Combined defects in oxidative phosphorylation and fatty acid β-oxidation in mitochondrial disease. Biosci Rep 36:e00313. - »Bioblast link«

Prochaska 2013 Springer CORRECTION.png
Ref. [27] Prochaska LJ, Cvetkov TL (2013) Mitochondrial electron transport. In: Roberts, G.C.K. (eds) Encyclopedia of Biophysics. Springer, Berlin, Heidelberg. - »Bioblast link«

Fisher-Wellman 2012 Trends Endocrinol Metab CORRECTION.png Fisher-Wellman 2012 Trends Endocrinol Metab Fig2 CORRECTION.png
Ref. [28] Fisher-Wellman KH, Neufer PD (2012) Linking mitochondrial bioenergetics to insulin resistance via redox biology. Trends Endocrinol Metab 23:142-53. - »Bioblast link«

Benard 2011 Springer CORRECTION.png
Ref. [29] Benard G, Bellance N, Jose C, Rossignol R (2011) Relationships between mitochondrial dynamics and bioenergetics. In: Lu Bingwei (ed) Mitochondrial dynamics and neurodegeneration. Springer ISBN 978-94-007-1290-4:47-68. - »Bioblast link«

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Ref. [30] Nussbaum RL (2005) Mining yeast in silico unearths a golden nugget for mitochondrial biology. J Clin Invest 115:2689-91. - »Bioblast link«

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Ref. [31] Sanchez H, Zoll J, Bigard X, Veksler V, Mettauer B, Lampert E, Lonsdorfer J, Ventura-Clapier R (2001) Effect of cyclosporin A and its vehicle on cardiac and skeletal muscle mitochondria: relationship to efficacy of the respiratory chain. Br J Pharmacol 133:781-8. - »Bioblast link«

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Ref. [32] Himms-Hagen J, Harper ME (2001) Physiological role of UCP3 may be export of fatty acids from mitochondria when fatty acid oxidation predominates: an hypothesis. Exp Biol Med (Maywood) 226:78-84. - »Bioblast link«

Brownlee 2001 Nature CORRECTION.png
Ref. [33] Brownlee M (2001) Biochemistry and molecular cell biology of diabetic complications. Nature 14:813-20. - »Bioblast link«
Ref. [34] Arden GB, Ramsey DJ (2015) Diabetic retinopathy and a novel treatment based on the biophysics of rod photoreceptors and dark adaptation. Webvision In: Kolb H, Fernandez E, Nelson R, eds. Webvision: The organization of the retina and visual system [Internet]. Salt Lake City (UT): University of Utah Health Sciences Center; 1995-. - »Bioblast link«

FADH2→CII misconceptions: Websites

The following graphs show zooms into the CII-related sections of figures found on the websites cited below. Erroneous presentations are marked by symbols.
OpenStax Biology.png
Website 1: OpenStax Biology - Fig. 7.10 Oxidative phosphorylation (CC BY 3.0). - OpenStax Biology got it wrong in figures and text. The error is copied without quality assessment and propagated in several links.
Website 2: Concepts of Biology - 1st Canadian Edition by Charles Molnar and Jane Gair - Fig. 4.19a
Website 3: LibreTexts Biology - Figure 7.11.1
Website 4: lumen Biology for Majors I - Fig. 1
Website 5: Pharmaguideline
Khan Academy modified from OpenStax CORRECTION.png
Website 6: Khan Academy - Image modified from "Oxidative phosphorylation: Figure 1", by OpenStax College, Biology (CC BY 3.0). Figure and text underscore the FADH2-error: "FADH2 .. feeds them (electrons) into the transport chain through complex II."
Website 7: Saylor Academy
Jack Westin CORRECTION.png
Website 8: Jack Westin MCAT Courses
Expii OpenStax CORRECTION.png
Website 1: OpenStax Biology - Fig. 7.12
Website 6: Khan Academy - Image modified from "Oxidative phosphorylation: Figure 3," by Openstax College, Biology (CC BY 3.0)
Website 7: Saylor Academy
Website 9: expii - Image source: By CNX OpenStax
Labxchange CORRECTION.png
Website 10: Labxchange - Figure 8.15 credit: modification of work by Klaus Hoffmeier CORRECTION.png
Website 4: lumen Biology for Majors I - Fig. 3
Website 9: expii - By OpenStax College CC BY 3.0, via Wikimedia Commons
Website 11: wikimedia 30148497 - Anatomy & Physiology, Connexions Web site., 2013-06-19
Website 12: 2018-08-21
Website 13: Quora
Website 14: TeachMePhysiology - Fig. 1. 2023-03-13
Website 15: ThoughtCo
Website 16: toppr
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Website 17: researchtweet
Website 18: Microbe Notes
BiochemDen CORRECTION.png
Website 19:
Vector Mine CORRECTION.png
Website 20: dreamstime
Website 21: VectorMine
Creative-biolabs CORRECTION.png
Website 22: creative-biolabs
Khan Academy CORRECTION.png
Website 6: Khan Academy
Website 7: Saylor Academy
Expii-Whitney, Rolfes 2002 CORRECTION.png
Website 9: expii - Whitney, Rolfes 2002
FlexBooks 2 0 CORRECTION.png
Website 23: FlexBooks - CK-12 Biology for High School- 2.28 Electron Transport, Figure 2
Hyperphysics CORRECTION.png
Website 24: hyperphysics
Labster Theory CORRECTION.png
Website 25: Labster Theory CORRECTION.png
Website 26:
Quizlet CORRECTION.png
Website 27: Quizlet
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Website 28: ScienceDirect
ScienceFacts CORRECTION.png
Website 29: ScienceFacts
Website 31:
Wikimedia ETC CORRECTION.png
Website 9: expii - By User:Rozzychan CC BY-SA 2.5, via Wikimedia Commons
Website 32: Wikimedia
YouTube Dirty Medicine Biochemistry CORRECTION.png
Website 33: YouTube Dirty Medicine Biochemistry - Uploaded 2019-07-18
YouTube sciencemusicvideos CORRECTION.png
Website 34: YouTube sciencemusicvideos - Uploaded 2014-08-19
ThoughtCo-Getty Images CORRECTION.png
Website 15: ThoughtCo - extender01 / iStock / Getty Images Plus
Website 17: dreamstime

CII and fatty acid oxidation

F-junction Wang 2019 Fig8.png
Fatty acid oxidation requires electron transferring flavoprotein CETF and CI for electron entry into the Q-junction (Gnaiger 2020; Wang et al 2019; see figures on the right).
Missaglia 2021 Crit Rev Biochem Mol Biol CORRECTION.png
When FADH2 is erroneously shown as a substrate of CII (1), a role of CII in fatty acid oxidation is suggested as a consequence (2).
Expii-Gabi Slizewska CORRECTION.png
Website 9: expii - Image source: By Gabi Slizewska
  • "Since mitochondrial Complex II also participates in the oxidation of fatty acids (6), .." (quote from Lemmi et al 1990).
  • Ref 6: Tzagoloff A (1982) Mitochondria. Plenum, New York.
  • This quote is erroneous, since the textbook by Tzagoloff (1982) represents fatty acid oxidation in figures and text without any involvement of CII.
FAO-CII Medical Biochemistry Page.jpg
Website 35: The Medical Biochemistry Page (accessed 2023-03-16)
Website 36: CHM333 LECTURES 37 & 38: 4/27 – 29/13 SPRING 2013 Professor Christine Hrycyna - Acyl-CoA dehydrogenase is listed under 'Electron transfer in Complex II'.
Website 37: Conduct Science: "In Complex II, the enzyme succinate dehydrogenase in the inner mitochondrial membrane reduce FADH2 to FAD+. Simultaneously, succinate, an intermediate in the Krebs cycle, is oxidized to fumarate." - Comments: FAD does not have a postive charge. FADH2 is the reduced form, it is not reduced. And again: In CII, FAD is reduced to FADH2.

MitoPedia concepts: MiP concept 

MitoPedia topics: Enzyme, Substrate and metabolite