NADH dehydrogenase (ubiquinone) flavoprotein 1，簡稱NDUFV1，為粒線體酵素複合體I中由細胞核基因體組表現的核心蛋白。此蛋白具有一個高度保留的黃素單核苷酸(FMN)的結合位置，可提供菸鹼醯胺腺嘌呤二核苷酸(NADH)氧化產生的電子進入粒線體的電子傳遞鏈；另外還含有一個高度保留的半胱胺酸的模組，可幫助形成一組[4Fe-4S]的N3鐵硫中心，而這個鐵硫中心在粒線體酵素複合體I的電子傳遞過程中扮演重要的角色。NDUFV1的缺失已經被發現和一些神經相關疾病例如心理性肌退化有關，而位於FMN結合位置的點突變也被發現存在於患有Leigh症候群的病人中。另一方面，粒線體酵素複合體I~V的組合對於粒線體能量產生的功能也是相當重要。此外，粒線體酵素複合體之間會形成更大型的複合體，用以增進電子傳遞的效率。目前由酵素複合體I、III和IV所形成的呼吸體酵素複合體(respirasome)已經被發現，且其缺失會造成腦肌病變及神經退化性疾病。
在本研究中，我們使用核酸干擾(RNAi)技術抑制人類胚胎腎細胞(HEK293)中NDUFV1的表現，藉此探討NDUFV1缺失後所帶來的影響。實驗結果顯示，NDUFV1受到抑制的細胞生長速度以及耗氧速率會下降，其粒線體酵素複合體I的功能亦有所降低。此外，NDUFV1的抑制也會使細胞內粒線體的膜電位降低，進而導致三磷酸腺苷(ATP)生成量的減少。同時在酵素複合體I的蛋白質分析中也發現NDUFV1的抑制會導致酵素複合體I的減少，同時也可能導致酵素複合體III和IV的減少。進一步的呼吸體酵素複合體分析顯示抑制NDUFV1也會使粒線體呼吸體酵素複合體減少，顯示著NDUFV1在調控粒線體酵素複合體的形成及穩定性上扮演著非常重要的角色。

NADH dehydrogenase (ubiquinone) flavoprotein 1 (NDUFV1) is a
nuclear-encoded subunit of mitochondrial Complex I. It provides a conserved FMN
binding site and is responsible for transferring electrons from NADH to FMN to
facilitate the entrance of electrons into the electron transport chain (ETC). NDUFV1
also has an iron sulfur cluster [4Fe-4S] (N3) that is the first of that kind in the
Complex I. The defect in NDUFV1 could cause neural disease like psychomotor
retardation, and some point mutations on FMN binding site have been found in
patients with Leigh syndrome. The assembly of intact OXPHOS complexes (complex
I~V) is very important in mitochondrial energy production. In addition, the assembly
of several respiratory complexes to form a larger supercomplex may improve the
efficiency of electron transport. The supercomplex composed by complex I, complex
III and complex IV has been confirmed and the defect in supercomplex assembly is
associated with encephalomyopathies and neurodegenerative disorders.
In present study, we applied a RNA interference system to suppress the
NDUFV1 expression in human embryonic kidney cell line (HEK293), and generated
three knockdown cell lines A9, B5 and E12 with 55~70% reduction in NDUFV1
protein level. We also performed oxygen consumption assay, dynamic complex I
activity assay, ATP determination assay and cell growth rate measurement to evaluate
the effects of NDUFV1 suppression. The results showed that the metabolic activity
and growth rate were significantly decreased in NDUFV1 knockdown cell lines. In
the respiratory complex assembly study, we solubilized mitochondria and applied a
high resolution clear native electrophoresis (HrCNE) approach to investigate whether
NDUFV1 knockdown affects individual respiratory complex assembly. The results
III
indicated that NDUFV1 knockdown would significantly decrease the level of complex I, III and IV. Further investigation of OXPHOS supercomplex formation also showed that while NDUFV1 was suppressed, CI/CIII2/CIVn supercomplexes were significantly reduced in cells. These finding suggests that NDUFV1 might play an important role in the assembly/stability of mitochondrial OXPHOS complexes and supercomplexes.

1. Murphy MP. How mitochondria produce reactive oxygen species. Biochem J. 2009 Jan 1;417(1):1-13.
2. Roland Lill. Function and biogenesis of iron–sulphur proteins. Nature. 2009 Aug 13;460(7257):831-8.
3. Chance Ba W, G. R. The respiratory chain and oxidative phosphorylation, in advances in enzymology and related areas of molecular biology volume 17. (ed F.F. Nord).
4. Rouslan G. Efremov, Rozbeh Baradaran, Leonid A. Sazanov. The architecture of respiratory complex I. Nature. 2010 May 27;465(7297):441-5.
5. DiMauro S, Schon EA. Mitochondrial respiratory-chain diseases. N Engl J Med. 2003 Jun 26;348(26):2656-68.
6. Munnich A, Rustin P, Rötig A, Chretien D, Bonnefont JP, Nuttin C, Cormier V, Vassault A, Parvy P, Bardet J, et al. Clinical aspects of mitochondrial disorders. J Inherit Metab Dis. 1992;15(4):448-55.
7. Hutchison CA 3rd, Newbold JE, Potter SS, Edgell MH. Maternal inheritance of mammalian mitochondrial DNA. Nature. 1974 Oct 11;251(5475):536-8.
8. Wallace DC. Mitotic segregation of mitochondrial DNAs in human cell hybrids and expression of chloramphenicol resistance. Somat Cell Mol Genet. 1986 Jan;12(1):41-9.
9. Ivanov PL, Wadhams MJ, Roby RK, Holland MM, Weedn VW, Parsons TJ. Mitochondrial DNA sequence heteroplasmy in the Grand Duke of Russia Georgij Romanov establishes the authenticity of the remains of Tsar Nicholas II. Nat Genet. 1996 Apr;12(4):417-20.
10. Holt IJ, Harding AE, Morgan-Hughes JA. Deletions of muscle mitochondrial DNA in patients with mitochondrial myopathies. Nature. 1988 Feb 25;331(6158):717-9.
11. Wallace DC, Singh G, Lott MT, Hodge JA, Schurr TG, Lezza AM, Elsas LJ 2nd, Nikoskelainen EK. Mitochondrial DNA mutation associated with Leber's hereditary optic neuropathy. Science. 1988 Dec 9;242(4884):1427-30.
60
12. Janssen RJ, van den Heuvel LP, Smeitink JA. Genetic defects in the oxidative phosphorylation (OXPHOS) system. Expert Rev Mol Diagn. 2004 Mar;4(2):143-56.
13. Hirst J. Mitochondrial complex I. Annu Rev Biochem. 2013;82:551-75.
14. Hofhaus G, Weiss H, Leonard K. Electron microscopic analysis of the peripheral and membrane parts of mitochondrial NADH dehydrogenase (complex I). J Mol Biol. 1991 Oct 5;221(3):1027-43.
15. Djafarzadeh R, Kerscher S, Zwicker K, Radermacher M, Lindahl M, Schägger H, Brandt U. Biophysical and structural characterization of proton-translocating NADH-dehydrogenase (complex I) from the strictly aerobic yeast Yarrowia lipolytica. Biochim Biophys Acta. 2000 Jul 20;1459(1):230-8.
16. Guénebaut V, Schlitt A, Weiss H, Leonard K, Friedrich T. Consistent structure between bacterial and mitochondrial NADH:ubiquinone oxidoreductase (complex I). J Mol Biol. 1998 Feb 13;276(1):105-12.
17. Grigorieff N. Three-dimensional structure of bovine NADH:ubiquinone oxidoreductase (complex I) at 22 A in ice. J Mol Biol. 1998 Apr 17;277(5):1033-46.
18. Friedrich T, Scheide D. The respiratory complex I of bacteria, archaea and eukarya and its module common with membrane-bound multisubunit hydrogenases. FEBS Lett. 2000 Aug 11;479(1-2):1-5.
19. Leif H, Weidner U, Berger A, Spehr V, Braun M, van Heek P, Friedrich T, Ohnishi T, Weiss H. Escherichia coli NADH dehydrogenase I, a minimal form of the mitochondrial complex I. Biochem Soc Trans. 1993 Nov;21(4):998-1001.
20. Hirst J, Carroll J, Fearnley IM, Shannon RJ, Walker JE. The nuclear encoded subunits of complex I from bovine heart mitochondria. Biochim Biophys Acta. 2003 Jul 10;1604(3):135-50.
21. Au HC, Seo BB, Matsuno-Yagi A, Yagi T, Scheffler IE. The NDUFA1 gene product (MWFE protein) is essential for activity of complex I in mammalian mitochondria. Proc Natl Acad Sci U S A. 1999 Apr 13;96(8):4354-9.
22. Kirby DM, Salemi R, Sugiana C, Ohtake A, Parry L, Bell KM, Kirk EP, Boneh A, Taylor RW, Dahl HH, Ryan MT, Thorburn DR. NDUFS6 mutations are a novel cause of lethal neonatal mitochondrial complex I deficiency. J Clin Invest. 2004 Sep;114(6):837-45.61
23. Hoefs SJ, van Spronsen FJ, Lenssen EW, Nijtmans LG, Rodenburg RJ, Smeitink JA, van den Heuvel LP. NDUFA10 mutations cause complex I deficiency in a patient with Leigh disease. Eur J Hum Genet. 2011 Mar;19(3):270-4.
24. Robinson, B.H. Human complex I deficiency: clinical spectrum and involvement of oxygen free radicals in the pathogenicity of the defect. Biochim Biophys Acta. 1998 May 6;1364(2):271-86.
25. Kirby DM, Crawford M, Cleary MA, Dahl HH, Dennett X, Thorburn DR. Respiratory chain complex I deficiency: an underdiagnosed energy generation disorder. Neurology. 1999 Apr 12;52(6):1255-64.
26. Loeffen JL, Smeitink JA, Trijbels JM, Janssen AJ, Triepels RH, Sengers RC, van den Heuvel LP. Isolated complex I deficiency in children: clinical, biochemical and genetic Hum Mutat. 2000;15(2):123-34.
27. Brandt U. Energy converting NADH:quinone oxidoreductase (complex I). Annu Rev Biochem. 2006;75:69-92.
28. Sazanov LA, Hinchliffe P. Structure of the hydrophilic domain of respiratory complex I from Thermus thermophilus. Science. 2006 Mar 10;311(5766):1430-6.
29. Efremov RG, Sazanov LA. Structure of the membrane domain of respiratory complex I. Nature. 2011 Aug 7;476(7361):414-20
30. Nouws J, Nijtmans LG, Smeitink JA, Vogel RO. Assembly factors as a new class of disease genes for mitochondrial complex I deficiency: cause, pathology and treatment options. Brain. 2012 Jan;135(Pt 1):12-22.
31. Videira A. Complex I from the fungus Neurospora crassa. Biochim Biophys Acta. 1998 May 6;1364(2):89-100.
32. Braun M, Bungert S, Friedrich T. Characterization of the overproduced NADH dehydrogenase fragment of the NADH:ubiquinone oxidoreductase (complex I) from Escherichia coli. Biochemistry. 1998 Feb 17;37(7):1861-7.
33. Kervinen M, Hinttala R, Helander HM, Kurki S, Uusimaa J, Finel M, Majamaa K, Hassinen IE. The MELAS mutations 3946 and 3949 perturb the critical structure in a conserved loop of the ND1 subunit of mitochondrial complex I. Hum Mol Genet. 2006 Sep 1;15(17):2543-52.
34. Kao MC, Di Bernardo S, Nakamaru-Ogiso E, Miyoshi H, Matsuno-Yagi A, Yagi T. Characterization of the membrane domain subunit NuoJ (ND6) of the NADH-quinone oxidoreductase from Escherichia coli by chromosomal DNA manipulation. Biochemistry. 2005 Mar 8;44(9):3562-71.
35. Kao MC, Nakamaru-Ogiso E, Matsuno-Yagi A, Yagi T. Characterization of the membrane domain subunit NuoK (ND4L) of the NADH-quinone oxidoreductase from Escherichia coli. Biochemistry. 2005 Jul 12;44(27):9545-54.
36. Holt PJ, Morgan DJ, Sazanov LA. The location of NuoL and NuoM subunits in the membrane domain of the Escherichia coli complex I: implications for the mechanism of proton pumping. J Biol Chem. 2003 Oct 31;278(44):43114-20.
37. Tuschen G, Sackmann U, Nehls U, Haiker H, Buse G, Weiss H. Assembly of NADH: ubiquinone reductase (complex I) in Neurospora mitochondria. Independent pathways of nuclear-encoded and mitochondrially encoded subunits. J Mol Biol. 1990 Jun 20;213(4):845-57.
38. Küffner R, Rohr A, Schmiede A, Krüll C, Schulte U. Involvement of two novel chaperones in the assembly of mitochondrial NADH:Ubiquinone oxidoreductase (complex I). J Mol Biol. 1998 Oct 23;283(2):409-17.
39. Schulte U, Fecke W, Krüll C, Nehls U, Schmiede A, Schneider R, Ohnishi T, Weiss H. In vivo dissection of the mitochondrial respiratory NADH: ubiquinone oxidoreductase (complex I). Biochim Biophys Acta. 1994 Aug 30;1187(2):121-4.
40. Hunte C, Zickermann V, Brandt U. Functional modules and structural basis of conformational coupling in mitochondrial complex I. Science. 2010 Jul 23;329(5990):448-51.
41. Cardol P, Lapaille M, Minet P, Franck F, Matagne RF, Remacle C. ND3 and ND4L subunits of mitochondrial complex I, both nucleus encoded in Chlamydomonas reinhardtii, are required for activity and assembly of the enzyme. Eukaryot Cell. 2006 Sep;5(9):1460-7.
42. Cardol P, Matagne RF, Remacle C. Impact of mutations affecting ND mitochondria-encoded subunits on the activity and assembly of complex I in Chlamydomonas. Implication for the structural organization of the enzyme. J Mol Biol. 2002 Jun 21;319(5):1211-21.
43. Remacle C, Duby F, Cardol P, Matagne RF. Mutations inactivating mitochondrial genes in Chlamydomonas reinhardtii. Biochem Soc Trans. 2001 Aug;29(Pt 4):442-6
44. Cardol P, Boutaffala L, Memmi S, Devreese B, Matagne RF, Remacle C. In Chlamydomonas, the loss of ND5 subunit prevents the assembly of whole mitochondrial complex I and leads to the formation of a low abundant 700 kDa subcomplex. Biochim Biophys Acta. 2008 Apr;1777(4):388-96.
45. Remacle C, Barbieri MR, Cardol P, Hamel PP. Eukaryotic complex I: functional diversity and experimental systems to unravel the assembly process. Mol Genet Genomics. 2008 Aug;280(2):93-110.
46. Perales-Clemente E, Fernández-Vizarra E, Acín-Pérez R, Movilla N, Bayona-Bafaluy MP, Moreno-Loshuertos R, Pérez-Martos A, Fernández-Silva P, Enríquez JA. Five entry points of the mitochondrially encoded subunits in mammalian complex I assembly. Mol Cell Biol. 2010 Jun;30(12):3038-47.
47. Scheffler IE, Yadava N, Potluri P. Molecular genetics of complex I-deficient Chinese hamster cell lines. Biochim Biophys Acta. 2004 Dec 6;1659(2-3):160-71.
48. Yadava N, Potluri P, Smith EN, Bisevac A, Scheffler IE. Species-specific and mutant MWFE proteins. Their effect on the assembly of a functional mammalian mitochondrial complex I. J Biol Chem. 2002 Jun 14;277(24):21221-30.
49. Yadava N, Houchens T, Potluri P, Scheffler IE. Development and characterization of a conditional mitochondrial complex I assembly system. J Biol Chem. 2004 Mar 26;279(13):12406-13.
50. Bourges I, Ramus C, Mousson de Camaret B, Beugnot R, Remacle C, Cardol P, Hofhaus G, Issartel JP. Structural organization of mitochondrial human complex I: role of the ND4 and ND5 mitochondria-encoded subunits and interaction with prohibitin. Biochem J. 2004 Nov 1;383(Pt. 3):491-9.
51. Ugalde C, Vogel R, Huijbens R, Van Den Heuvel B, Smeitink J, Nijtmans L. Human mitochondrial complex I assembles through the combination of evolutionary conserved modules: a framework to interpret complex I deficiencies. Hum Mol Genet. 2004 Oct 15;13(20):2461-72.
52. Vogel RO, Dieteren CE, van den Heuvel LP, Willems PH, Smeitink JA, Koopman WJ, Nijtmans LG. Identification of mitochondrial complex I assembly intermediates by tracing tagged NDUFS3 demonstrates the entry point of mitochondrial subunits. J Biol Chem. 2007 Mar 9;282(10):7582-90.
53. Lazarou M, McKenzie M, Ohtake A, Thorburn DR, Ryan MT. Analysis of the assembly profiles for mitochondrial- and nuclear-DNA-encoded subunits into complex I. Mol Cell Biol. 2007 Jun;27(12):4228-37.
54. Lazarou M, Thorburn DR, Ryan MT, McKenzie M. Assembly of mitochondrial complex I and defects in disease. Biochim Biophys Acta. 2009 Jan;1793(1):78-88.
55. Mimaki M, Wang X, McKenzie M, Thorburn DR, Ryan MT. Understanding mitochondrial complex I assembly in health and disease. Biochim Biophys Acta. 2012 Jun;1817(6):851-62.
56. Cristina Ugalde, Rolf J.R.J. Janssen, Lambert P. van den Heuvel, Jan A.M. Smeitink and Leo G.J. Nijtmans. Differences in assembly or stability of complex I and other mitochondrial OXPHOS complexes in inherited complex I deficiency. Human Molecular Genetics, 2004, Vol.13, No.6, 659–667
57. Hackenbrock CR, Chazotte B, Gupte SS. The random collision model and a critical assessment of diffusion and collision in mitochondrial electron transport. J Bioenerg Biomembr. 1986 Oct;18(5):331-68.
58. Schägger H, Pfeiffer K. Supercomplexes in the respiratory chains of yeast and mammalian mitochondria. EMBO J. 2000 Apr 17;19(8):1777-83.
59. Acín-Pérez R, Fernández-Silva P, Peleato ML, Pérez-Martos A, Enriquez JA. Respiratory active mitochondrial supercomplexes. Mol Cell. 2008 Nov 21;32(4):529-39.
60. Lenaz G, Genova ML. Structure and organization of mitochondrial respiratory complexes: a new understanding of an old subject. Antioxid Redox Signal. 2010 Apr 15;12(8):961-1008.
61. Althoff T, Mills DJ, Popot JL, Kühlbrandt W. Arrangement of electron transport chain components in bovine mitochondrial supercomplex I1III2IV1. EMBO J. 2011 Sep 9;30(22):4652-64.
62. Dudkina NV, Kudryashev M, Stahlberg H, Boekema EJ. Interaction of complexes I, III, and IV within the bovine respirasome by single particle cryoelectron tomography. Proc Natl Acad Sci U S A. 2011 Sep 13;108(37):15196-200.
63. Heinemeyer J, Braun HP, Boekema EJ, Kouril R. A structural model of the cytochrome C reductase/oxidase supercomplex from yeast mitochondria. J Biol Chem. 2007 Apr 20;282(16):12240-8.
64. Schäfer E, Seelert H, Reifschneider NH, Krause F, Dencher NA, Vonck J. Architecture of active mammalian respiratory chain supercomplexes. J Biol Chem. 2006 Jun 2;281(22):15370-5.
65. Schägger H, Pfeiffer K. The ratio of oxidative phosphorylation complexes I-V in bovine heart mitochondria and the composition of respiratory chain supercomplexes. J Biol Chem. 2001 Oct 12;276(41):37861-7.
66. Moreno-Lastres D, Fontanesi F, García-Consuegra I, Martín MA, Arenas J, Barrientos A, Ugalde C. Mitochondrial complex I plays an essential role in human respirasome assembly. Cell Metab. 2012 Mar 7;15(3):324-35.
67. D'Aurelio M, Gajewski CD, Lenaz G, Manfredi G. Respiratory chain supercomplexes set the threshold for respiration defects in human mtDNA mutant cybrids. Hum Mol Genet. 2006 Jul 1;15(13):2157-69.
68. Fernandez-Vizarra E, Bugiani M, Goffrini P, Carrara F, Farina L, Procopio E, Donati A, Uziel G, Ferrero I, Zeviani M. Impaired complex III assembly associated with BCS1L gene mutations in isolated mitochondrial encephalopathy. Hum Mol Genet. 2007 May 15;16(10):1241-52.
69. Morán M, Marín-Buera L, Gil-Borlado MC, Rivera H, Blázquez A, Seneca S, Vázquez-López M, Arenas J, Martín MA, Ugalde C. Cellular pathophysiological consequences of BCS1L mutations in mitochondrial complex III enzyme deficiency. Hum Mutat. 2010 Aug;31(8):930-41.
70. Spencer SR, Taylor JB, Cowell IG, Xia CL, Pemble SE, Ketterer B. The human mitochondrial NADH: ubiquinone oxidoreductase 51-kDa subunit maps adjacent to the glutathione S-transferase P1-1 gene on chromosome 11q13. Genomics. 1992 Dec;14(4):1116-8.
71. de Coo RF, Buddiger PA, Smeets HJ, van Oost BA. The structure of the human NDUFV1 gene encoding the 51-kDa subunit of mitochondrial complex I. Mamm Genome. 1999 Jan;10(1):49-53.
72. Patel SD, Aebersold R, Attardi G. cDNA-derived amino acid sequence of the NADH-binding 51-kDa subunit of the bovine respiratory NADH dehydrogenase reveals striking similarities to a bacterial NAD(+)-reducing hydrogenase. Proc Natl Acad Sci U S A. 1991 May 15;88(10):4225-9.
73. Krishnamoorthy G, Hinkle PC. Studies on the electron transfer pathway, topography of iron-sulfur centers, and site of coupling in NADH-Q oxidoreductase. J Biol Chem. 1988 Nov 25;263(33):17566-75.
74. Velazquez I, Nakamaru-Ogiso E, Yano T, Ohnishi T, Yagi T. Amino acid residues associated with cluster N3 in the NuoF subunit of the proton-translocating NADH-quinone oxidoreductase from Escherichia coli. FEBS Lett. 2005 Jun 6;579(14):3164-8.
75. Euro L, Belevich G, Bloch DA, Verkhovsky MI, Wikström M, Verkhovskaya M. The role of the invariant glutamate 95 in the catalytic site of Complex I from Escherichia coli. Biochim Biophys Acta. 2009 Jan;1787(1):68-73.
76. Knuuti J, Belevich G, Sharma V, Bloch DA, Verkhovskaya M. A single amino acid residue controls ROS production in the respiratory Complex I from Escherichia coli. Mol Microbiol. 2013 Dec;90(6):1190-200.
77. Auriol C, Bestel-Corre G, Claude JB, Soucaille P, Meynial-Salles I. Stress-induced evolution of Escherichia coli points to original concepts in respiratory cofactor selectivity. Proc Natl Acad Sci U S A. 2011 Jan 25;108(4):1278-83.
78. Bénit P, Chretien D, Kadhom N, de Lonlay-Debeney P, Cormier-Daire V, Cabral A, Peudenier S, Rustin P, Munnich A, Rötig A. Large-scale deletion and point mutations of the nuclear NDUFV1 and NDUFS1 genes in mitochondrial complex I deficiency. Am J Hum Genet. 2001 Jun;68(6):1344-52.
79. Fearnley IM, Walker JE. Conservation of sequences of subunits of mitochondrial complex I and their relationships with other proteins. Biochim Biophys Acta. 1992 Dec 7;1140(2):105-34.
80. Marin SE, Mesterman R, Robinson B, Rodenburg RJ, Smeitink J, Tarnopolsky MA. Leigh syndrome associated with mitochondrial complex I deficiency due to novel mutations In NDUFV1 and NDUFS2. Gene. 2013 Mar 1;516(1):162-7.
81. Vilain C, Rens C, Aeby A, Balériaux D, Van Bogaert P, Remiche G, Smet J, Van Coster R, Abramowicz M, Pirson I. A novel NDUFV1 gene mutation in complex I deficiency in consanguineous siblings with brainstem lesions and Leigh syndrome. Clin Genet. 2012 Sep;82(3):264-70.
82. Genova ML, Castelluccio C, Fato R, Parenti Castelli G, Merlo Pich M, Formiggini G, Bovina C, Marchetti M, Lenaz G. Major changes in complex I activity in mitochondria from aged rats may not be detected by direct assay of NADH:coenzyme Q reductase. Biochem J. 1995 Oct 1;311 ( Pt 1):105-9.
83. Chen LB. Mitochondrial membrane potential in living cells. Annu Rev Cell Biol. 1988;4:155-81.
84. Nicholls DG. Mitochondrial membrane potential and aging. Aging Cell. 2004 Feb;3(1):35-40.
85. Wallace DC. A mitochondrial paradigm of metabolic and degenerative diseases, aging, and cancer: a dawn for evolutionary medicine. Annu Rev Genet. 2005;39:359-407.
86. Sazanov LA. Respiratory complex I: mechanistic and structural insights provided by the crystal structure of the hydrophilic domain. Biochemistry. 2007 Mar 6;46(9):2275-88.
87. Prüss BM, Nelms JM, Park C, Wolfe AJ. Mutations in NADH:ubiquinone oxidoreductase of Escherichia coli affect growth on mixed amino acids. J Bacteriol. 1994 Apr;176(8):2143-50.
88. Procaccio V, Depetris D, Soularue P, Mattei MG, Lunardi J, Issartel JP. cDNA sequence and chromosomal localization of the NDUFS8 human gene coding for the 23 kDa subunit of the mitochondrial complex I. Biochim Biophys Acta. 1997 Mar 20;1351(1-2):37-41.
89. Rasmussen T, Scheide D, Brors B, Kintscher L, Weiss H, Friedrich T. Identification of two tetranuclear FeS clusters on the ferredoxin-type subunit of NADH:ubiquinone oxidoreductase (complex I). Biochemistry. 2001 May 22;40(20):6124-31.
90. Rak M, Rustin P. Supernumerary subunits NDUFA3, NDUFA5 and NDUFA12 are required for the formation of the extramembrane arm of human mitochondrial complex I. FEBS Lett. 2014 May 2;588(9):1832-8.