Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Structural insight into the type-II mitochondrial NADH dehydrogenases


The single-component type-II NADH dehydrogenases (NDH-2s) serve as alternatives to the multisubunit respiratory complex I (type-I NADH dehydrogenase (NDH-1), also called NADH:ubiquinone oxidoreductase; EC in catalysing electron transfer from NADH to ubiquinone in the mitochondrial respiratory chain1. The yeast NDH-2 (Ndi1) oxidizes NADH on the matrix side and reduces ubiquinone to maintain mitochondrial NADH/NAD+ homeostasis. Ndi1 is a potential therapeutic agent for human diseases caused by complex I defects2,3,4,5,6,7,8,9, particularly Parkinson’s disease, because its expression restores the mitochondrial activity in animals with complex I deficiency. NDH-2s in pathogenic microorganisms are viable targets for new antibiotics10,11. Here we solve the crystal structures of Ndi1 in its substrate-free, NADH-, ubiquinone- and NADH–ubiquinone-bound states, to help understand the catalytic mechanism of NDH-2s. We find that Ndi1 homodimerization through its carboxy-terminal domain is critical for its catalytic activity and membrane targeting. The structures reveal two ubiquinone-binding sites (UQI and UQII) in Ndi1. NADH and UQI can bind to Ndi1 simultaneously to form a substrate–protein complex. We propose that UQI interacts with FAD to act as an intermediate for electron transfer, and that NADH transfers electrons through this FAD–UQI complex to UQII. Together our data reveal the regulatory and catalytic mechanisms of Ndi1 and may facilitate the development or targeting of NDH-2s for potential therapeutic applications.

Your institute does not have access to this article

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Ndi1 has a unique CTD.
Figure 2: The CTD mediates Ndi1 homodimerization and membrane attachment.
Figure 3: Electron transfer of Ndi1 involves two ubiquinone molecules.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The atomic coordinates and structure factors of the apo-, NADH-, ubiquinone- and NADH–ubiquinone-bound forms of Ndi1 have been deposited in the Protein Data Bank under accession codes 4G6G, 4G6H, 4G74 and 4G73, respectively.


  1. Melo, A. M., Bandeiras, T. M. & Teixeira, M. New insights into type II NAD(P)H:quinone oxidoreductases. Microbiol. Mol. Biol. Rev. 68, 603–616 (2004)

    CAS  PubMed  PubMed Central  Google Scholar 

  2. Sanz, A. et al. Expression of the yeast NADH dehydrogenase Ndi1 in Drosophila confers increased lifespan independently of dietary restriction. Proc. Natl Acad. Sci. USA 107, 9105–9110 (2010)

    CAS  PubMed  ADS  Google Scholar 

  3. Maas, M. F., Sellem, C. H., Krause, F., Dencher, N. A. & Sainsard-Chanet, A. Molecular gene therapy: overexpression of the alternative NADH dehydrogenase NDI1 restores overall physiology in a fungal model of respiratory complex I deficiency. J. Mol. Biol. 399, 31–40 (2010)

    CAS  PubMed  Google Scholar 

  4. Perales-Clemente, E. et al. Restoration of electron transport without proton pumping in mammalian mitochondria. Proc. Natl Acad. Sci. USA 105, 18735–18739 (2008)

    CAS  PubMed  ADS  Google Scholar 

  5. Marella, M., Seo, B. B., Flotte, T. R., Matsuno-Yagi, A. & Yagi, T. No immune responses by the expression of the yeast Ndi1 protein in rats. PLoS ONE 6, e25910 (2011)

    CAS  PubMed  PubMed Central  ADS  Google Scholar 

  6. Marella, M., Seo, B. B., Yagi, T. & Matsuno-Yagi, A. Parkinson’s disease and mitochondrial complex I: a perspective on the Ndi1 therapy. J. Bioenerg. Biomembr. 41, 493–497 (2009)

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Marella, M., Seo, B. B., Thomas, B. B., Matsuno-Yagi, A. & Yagi, T. Successful amelioration of mitochondrial optic neuropathy using the yeast NDI1 gene in a rat animal model. PLoS ONE 5, e11472 (2010)

    PubMed  PubMed Central  ADS  Google Scholar 

  8. Barber-Singh, J., Seo, B. B., Matsuno-Yagi, A. & Yagi, T. Protective role of rAAV-NDI1, serotype 5, in an acute MPTP mouse Parkinson’s model. Parkinson’s Dis. 2011, 438370 (2011)

    Google Scholar 

  9. Yagi, T. et al. Can a single subunit yeast NADH dehydrogenase (Ndi1) remedy diseases caused by respiratory complex I defects? Rejuvenation Res. 9, 191–197 (2006)

    CAS  PubMed  Google Scholar 

  10. Teh, J. S., Yano, T. & Rubin, H. Type II NADH: menaquinone oxidoreductase of Mycobacterium tuberculosis. Infect. Disord. Drug Targets 7, 169–181 (2007)

    CAS  PubMed  ADS  Google Scholar 

  11. Biagini, G. A. et al. Generation of quinolone antimalarials targeting the Plasmodium falciparum mitochondrial respiratory chain for the treatment and prophylaxis of malaria. Proc. Natl Acad. Sci. USA 109, 8298–8303 (2012)

    CAS  PubMed  ADS  Google Scholar 

  12. Rich, P. R. & Marechal, A. The mitochondrial respiratory chain. Essays Biochem. 47, 1–23 (2010)

    CAS  PubMed  Google Scholar 

  13. Efremov, R. G. & Sazanov, L. A. Structure of the membrane domain of respiratory complex I. Nature 476, 414–420 (2011)

    CAS  PubMed  ADS  Google Scholar 

  14. Walker, J. E. The NADH:ubiquinone oxidoreductase (complex I) of respiratory chains. Q. Rev. Biophys. 25, 253–324 (1992)

    CAS  PubMed  Google Scholar 

  15. Kerscher, S., Drose, S., Zickermann, V. & Brandt, U. The three families of respiratory NADH dehydrogenases. Results Probl. Cell Differ. 45, 185–222 (2008)

    CAS  PubMed  Google Scholar 

  16. Barquera, B., Zhou, W., Morgan, J. E. & Gennis, R. B. Riboflavin is a component of the Na+-pumping NADH-quinone oxidoreductase from Vibrio cholerae. Proc. Natl Acad. Sci. USA 99, 10322–10324 (2002)

    CAS  PubMed  ADS  Google Scholar 

  17. Ohnishi, T., Kawaguchi, K. & Hagihara, B. Preparation and some properties of yeast mitochondria. J. Biol. Chem. 241, 1797–1806 (1966)

    CAS  PubMed  Google Scholar 

  18. Ohnishi, T., Sottocasa, G. & Ernster, L. Current approaches to the mechanism of energy-coupling in the respiratory chain. Studies with yeast mitochondria. Bull. Soc. Chim. Biol. (Paris) 48, 1189–1203 (1966)

    CAS  Google Scholar 

  19. Kerscher, S. J. Diversity and origin of alternative NADH:ubiquinone oxidoreductases. Biochim. Biophys. Acta 1459, 274–283 (2000)

    CAS  PubMed  Google Scholar 

  20. Seo, B. B. et al. Molecular remedy of complex I defects: rotenone-insensitive internal NADH-quinone oxidoreductase of Saccharomyces cerevisiae mitochondria restores the NADH oxidase activity of complex I-deficient mammalian cells. Proc. Natl Acad. Sci. USA 95, 9167–9171 (1998)

    CAS  PubMed  ADS  Google Scholar 

  21. Holm, L. & Rosenstrom, P. Dali server: conservation mapping in 3D. Nucleic Acids Res. 38, W545–W549 (2010)

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Villegas, J. M., Volentini, S. I., Rintoul, M. R. & Rapisarda, V. A. Amphipathic C-terminal region of Escherichia coli NADH dehydrogenase-2 mediates membrane localization. Arch. Biochem. Biophys. 505, 155–159 (2011)

    CAS  PubMed  Google Scholar 

  23. Yang, Y. et al. Reaction mechanism of single subunit NADH-ubiquinone oxidoreductase (Ndi1) from Saccharomyces cerevisiae: evidence for a ternary complex mechanism. J. Biol. Chem. 286, 9287–9297 (2011)

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Yamashita, T., Nakamaru-Ogiso, E., Miyoshi, H., Matsuno-Yagi, A. & Yagi, T. Roles of bound quinone in the single subunit NADH-quinone oxidoreductase (Ndi1) from Saccharomyces cerevisiae. J. Biol. Chem. 282, 6012–6020 (2007)

    CAS  PubMed  Google Scholar 

  25. Murai, M. et al. Characterization of the ubiquinone binding site in the alternative NADH-quinone oxidoreductase of Saccharomyces cerevisiae by photoaffinity labeling. Biochemistry 49, 2973–2980 (2010)

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Sevrioukova, I. F., Poulos, T. L. & Churbanova, I. Y. Crystal structure of the putidaredoxin reductase x putidaredoxin electron transfer complex. J. Biol. Chem. 285, 13616–13620 (2010)

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Ingledew, W. J., Ohnishi, T. & Salerno, J. C. Studies on a stabilisation of ubisemiquinone by Escherichia coli quinol oxidase, cytochrome bo. Eur. J. Biochem. 227, 903–908 (1995)

    CAS  PubMed  Google Scholar 

  28. Adelroth, P., Paddock, M. L., Sagle, L. B., Feher, G. & Okamura, M. Y. Identification of the proton pathway in bacterial reaction centers: both protons associated with reduction of QB to QBH2 share a common entry point. Proc. Natl Acad. Sci. USA 97, 13086–13091 (2000)

    CAS  PubMed  ADS  Google Scholar 

  29. Okamura, M. Y., Paddock, M. L., Graige, M. S. & Feher, G. Proton and electron transfer in bacterial reaction centers. Biochim. Biophys. Acta 1458, 148–163 (2000)

    CAS  PubMed  Google Scholar 

  30. Ohnishi, T., Ohnishi, S. T., Shinzawa-Itoh, K., Yoshikawa, S. & Weber, R. T. EPR detection of two protein-associated ubiquinone components (SQ(Nf) and SQ(Ns)) in the membrane in situ and in proteoliposomes of isolated bovine heart complex I. Biochim. Biophys. Acta 1817, 1803–1809 (2012)

    CAS  PubMed  Google Scholar 

  31. Edgar, R. C. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res. 32, 1792–1797 (2004)

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Hall, T. A. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp. Ser. 41, 95–98 (1999)

    CAS  Google Scholar 

  33. Guindon, S. E. & Gascuel, O. A. Simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst. Biol. 52, 696–704 (2003)

    PubMed  Google Scholar 

  34. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997)

    CAS  PubMed  Google Scholar 

  35. Collaborative Computational Project. Number 4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D 50, 760–763 (1994)

  36. Schneider, T. R. & Sheldrick, G. M. Substructure solution with SHELXD. Acta Crystallogr. D 58, 1772–1779 (2002)

    PubMed  Google Scholar 

  37. Terwilliger, T. C. Rapid automatic NCS identification using heavy-atom substructures. Acta Crystallogr. D 58, 2213–2215 (2002)

    PubMed  Google Scholar 

  38. McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007)

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Cowtan, K. DM: an automated procedure for phase improvement by density modification. CCP4 Newslett. 31, 34–38 (1994)

    Google Scholar 

  40. Cowtan, K. The Buccaneer software for automated model building. 1. Tracing protein chains. Acta Crystallogr. D 62, 1002–1011 (2006)

    PubMed  Google Scholar 

  41. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D 60, 2126–2132 (2004)

    PubMed  Google Scholar 

  42. Adams, P. D. et al. PHENIX: building new software for automated crystallographic structure determination. Acta Crystallogr. D 58, 1948–1954 (2002)

    MathSciNet  PubMed  Google Scholar 

Download references


We would like to thank the staff at beamline BL17U of the Shanghai Synchrotron Radiation Facility for their assistance with data collection. We thank W. Tong for maintenance and support of the EPR facility at the High Magnetic Field Laboratory, Chinese Academy of Sciences. This work was supported by the Ministry of Science and Technology of China (2011CB910502, 2011CB910900 and 2012CB911101), the National Natural Science Foundation of China (31030020 and 31170679) and Chinese Key Research Plan-Protein Sciences (2011CB911104).

Author information

Authors and Affiliations



M.Y. designed and directed the project. Y.F., W.L., J. W., J.G., D.X. and J.-W.W. purified the proteins, grew the crystals, collected data, solved the crystal structures and performed the in vitro activity analyses. Y.L. and Q.Z. performed the genome analysis of the NDHs. B.Z. and J.L. performed the in vivo biological analyses. Y.F., K.W. and C.T. performed the EPR analyses. M.Y. analysed the data and wrote the paper with the help of all the authors.

Corresponding author

Correspondence to Maojun Yang.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-13 and Supplementary Tables 1-2. (PDF 4938 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Feng, Y., Li, W., Li, J. et al. Structural insight into the type-II mitochondrial NADH dehydrogenases. Nature 491, 478–482 (2012).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing