Structural Bioenergetics
Proton pumping NADH:ubiquinone oxidoreductase (complex I) is a very large membrane protein complex with a key function for efficient energy production of the cell. The minimal form of complex I found in bacteria comprises 14 conserved subunits. The larger and more complicated enzyme of the mitochondrial respiratory chain harbors more than 40 subunits with a total mass of almost 1 MDa.
Complex I utilizes the redox energy released in the electron transfer reaction from NADH to ubiquinone to pump protons across the inner mitochondrial membrane or the cell membrane of bacteria. Redox linked proton transloaction by complex I contributes significantly to the electrochemical proton gradient that ultimately drives ATP synthesis in oxidative phosphorylation. A variety of neuromuscular and neurodegenerative human diseases has been linked with complex I dysfunction. However, the molecular details of energy conversion by complex I are still poorly understood. We use the aerobic yeast Yarrowia lipolytica as a yeast genetic model system to study eukaryotic complex I. Protein purification by His-tag affinity purification and genetic manipulation of nuclear encoded subunits is straightforward. We have accomplished the first X-ray crystallographic analysis of mitochondrial complex I. The structure provided exciting insight into the molecular basis of redox-driven proton pumping. Our work is supported by the Deutsche Forschungsgemeinschaft (ZI 552/3-1, ZI 552 4-1 and EXC 115);and our goal is to obtain a comprehensive understanding of complex I, a giant molecular machine.
Dr. Heike Angerer
Karin Siegmund
Prof. Volker Zickermann
Galemou Yoga E, Parey K, Djurabekova A, Haapanen O, Siegmund K, Zwicker K, Sharma V, Zickermann V, Angerer H Essential role of accessory subunit LYRM6 in the mechanism of mitochondrial complex I. Nat Commun 2020. 11 (1) 6008 Link
Scherr J, Tang Z, Küllmer M, Balser S, Scholz AS, Winter A, Parey K, Rittner A, Grininger M, Zickermann V, Rhinow D, Terfort A, Turchanin A Smart Molecular Nanosheets for Advanced Preparation of Biological Samples in Electron Cryo-Microscopy. ACS Nano 2020. 14 (8) 9972-9978 Link
Parey K, Wirth C, Vonck J, Zickermann V Respiratory complex I - structure, mechanism and evolution. Curr Opin Struct Biol 2020. 63 1-9 Link
Galemou Yoga E, Angerer H, Parey K, Zickermann V Respiratory complex I - Mechanistic insights and advances in structure determination. Biochim Biophys Acta Bioenerg 2020. 1861 (3) 148153 Link
Parey K, Haapanen O, Sharma V, Köfeler H, Züllig T, Prinz S, Siegmund K, Wittig I, Mills DJ, Vonck J, Kühlbrandt W, Zickermann V High-resolution cryo-EM structures of respiratory complex I: Mechanism, assembly, and disease. Sci Adv 2019. 5 (12) eaax9484 Link
Galemou Yoga E, Haapanen O, Wittig I, Siegmund K, Sharma V, Zickermann V Mutations in a conserved loop in the PSST subunit of respiratory complex I affect ubiquinone binding and dynamics. Biochim Biophys Acta Bioenerg 2019. 1860 (7) 573-581 Link
Scherr J, Neuhaus A, Parey K, Klusch N, Murphy BJ, Zickermann V, Kühlbrandt W, Terfort A, Rhinow D Noncovalent Functionalization of Carbon Substrates with Hydrogels Improves Structural Analysis of Vitrified Proteins by Electron Cryo-Microscopy. ACS Nano 2019. 13 (6) 7185-7190 Link
Cabrera-Orefice A, Yoga EG, Wirth C, Siegmund K, Zwicker K, Guerrero-Castillo S, Zickermann V, Hunte C, Brandt U Locking loop movement in the ubiquinone pocket of complex I disengages the proton pumps. Nat Commun 2018. 9 (1) 4500 Link
Parey K, Brandt U, Xie H, Mills DJ, Siegmund K, Vonck J, Kühlbrandt W, Zickermann V Cryo-EM structure of respiratory complex I at work. Elife 2018. 7 Link
Angerer H, Schönborn S, Gorka J, Bahr U, Karas M, Wittig I, Heidler J, Hoffmann J, Morgner N, Zickermann V Acyl modification and binding of mitochondrial ACP to multiprotein complexes. Biochim Biophys Acta Mol Cell Res 2017. 1864 (10) 1913-1920 Link
Scherr J, Parey K, Klusch N, Murphy BJ, Balser S, Neuhaus A, Zickermann V, Kühlbrandt W, Terfort A, Rhinow D Self-Perforated Hydrogel Nanomembranes Facilitate Structural Analysis of Proteins by Electron Cryo-Microscopy. ACS Nano 2017. 11 (6) 6467-6473 Link
Krishnathas R, Bonke E, Dröse S, Zickermann V, Nasiri HR Identification of 4--[2-(4-phenoxyphenyl)ethyl]quinazoline-4,6-diamine as a novel, highly potent and specific inhibitor of mitochondrial complex I. Medchemcomm 2017. 8 (3) 657-661 Link
Kahlhöfer F, Kmita K, Wittig I, Zwicker K, Zickermann V Accessory subunit NUYM (NDUFS4) is required for stability of the electron input module and activity of mitochondrial complex I. Biochim Biophys Acta Bioenerg 2017. 1858 (2) 175-181 Link
D'Imprima E, Mills DJ, Parey K, Brandt U, Kühlbrandt W, Zickermann V, Vonck J Cryo-EM structure of respiratory complex I reveals a link to mitochondrial sulfur metabolism. Biochim Biophys Acta 2016. 1857 (12) 1935-1942 Link
Hahn A, Parey K, Bublitz M, Mills DJ, Zickermann V, Vonck J, Kühlbrandt W, Meier T Structure of a Complete ATP Synthase Dimer Reveals the Molecular Basis of Inner Mitochondrial Membrane Morphology. Mol Cell 2016. 63 (3) 445-56 Link
Brandt UU, Zickermann V Preface to complex I special issue. Biochim Biophys Acta 2016. 1857 (7) 861-2 Link
Wirth C, Brandt U, Hunte C, Zickermann V Structure and function of mitochondrial complex I. Biochim Biophys Acta 2016. 1857 (7) 902-14 Link
Kmita K, Wirth C, Warnau J, Guerrero-Castillo S, Hunte C, Hummer G, Kaila VR, Zwicker K, Brandt U, Zickermann V Accessory NUMM (NDUFS6) subunit harbors a Zn-binding site and is essential for biogenesis of mitochondrial complex I. Proc Natl Acad Sci U S A 2015. 112 (18) 5685-90 Link
Zickermann V, Wirth C, Nasiri H, Siegmund K, Schwalbe H, Hunte C, Brandt U Structural biology. Mechanistic insight from the crystal structure of mitochondrial complex I. Science 2015. 347 (6217) 44-9 Link
Angerer H, Radermacher M, Mańkowska M, Steger M, Zwicker K, Heide H, Wittig I, Brandt U, Zickermann V The LYR protein subunit NB4M/NDUFA6 of mitochondrial complex I anchors an acyl carrier protein and is essential for catalytic activity. Proc Natl Acad Sci U S A 2014. 111 (14) 5207-12 Link
Kmita K, Zickermann V Accessory subunits of mitochondrial complex I. Biochem Soc Trans 2013. 41 (5) 1272-9 Link
Radermacher M, Ruiz T, Fowler DJ, Yu L, Dröse S, Krack S, Kerscher S, Zickermann V, Brandt U 3D Reconstruction of a Subcomplex of NADH-ubiquinone-oxidoreductase (Complex I) from Yarrowia lipolytica. Microsc Microanal 2011. 17 (S2) 90-91 Link
Ruzzenente B, Metodiev MD, Wredenberg A, Bratic A, Park CB, Cámara Y, Milenkovic D, Zickermann V, Wibom R, Hultenby K, Erdjument-Bromage H, Tempst P, Brandt U, Stewart JB, Gustafsson CM, Larsson NG LRPPRC is necessary for polyadenylation and coordination of translation of mitochondrial mRNAs. EMBO J 2012. 31 (2) 443-56 Link
Dröse S, Krack S, Sokolova L, Zwicker K, Barth HD, Morgner N, Heide H, Steger M, Nübel E, Zickermann V, Kerscher S, Brutschy B, Radermacher M, Brandt U Functional dissection of the proton pumping modules of mitochondrial complex I. PLoS Biol 2011. 9 (8) e1001128 Link
Davies KM, Strauss M, Daum B, Kief JH, Osiewacz HD, Rycovska A, Zickermann V, Kühlbrandt W Macromolecular organization of ATP synthase and complex I in whole mitochondria. Proc Natl Acad Sci U S A 2011. 108 (34) 14121-6 Link
Angerer H, Zwicker K, Wumaier Z, Sokolova L, Heide H, Steger M, Kaiser S, Nübel E, Brutschy B, Radermacher M, Brandt U, Zickermann V A scaffold of accessory subunits links the peripheral arm and the distal proton-pumping module of mitochondrial complex I. Biochem J 2011. 437 (2) 279-88 Link
Ladig R, Sommer MS, Hahn A, Leisegang MS, Papasotiriou DG, Ibrahim M, Elkehal R, Karas M, Zickermann V, Gutensohn M, Brandt U, Klösgen RB, Schleiff E A high-definition native polyacrylamide gel electrophoresis system for the analysis of membrane complexes. Plant J 2011. 67 (1) 181-94 Link
Hunte C, Zickermann V, Brandt U Functional modules and structural basis of conformational coupling in mitochondrial complex I. Science 2010. 329 (5990) 448-51 Link
Tocilescu MA, Zickermann V, Zwicker K, Brandt U Quinone binding and reduction by respiratory complex I. Biochim Biophys Acta 2010. 1797 (12) 1883-90 Link
Zickermann V, Angerer H, Ding MG, Nübel E, Brandt U Small single transmembrane domain (STMD) proteins organize the hydrophobic subunits of large membrane protein complexes. FEBS Lett 2010. 584 (12) 2516-25 Link
Zickermann V, Wumaier Z, Wrzesniewska B, Hunte C, Schägger H Native immunoblotting of blue native gels to identify conformation-specific antibodies. Proteomics 2010. 10 (1) 159-63 Link
Clason T, Ruiz T, Schägger H, Peng G, Zickermann V, Brandt U, Michel H, Radermacher M The structure of eukaryotic and prokaryotic complex I. J Struct Biol 2010. 169 (1) 81-8 Link
Zickermann V, Kerscher S, Zwicker K, Tocilescu MA, Radermacher M, Brandt U Architecture of complex I and its implications for electron transfer and proton pumping. Biochim Biophys Acta 2009. 1787 (6) 574-83 Link
Zickermann V, Dröse S, Tocilescu MA, Zwicker K, Kerscher S, Brandt U Challenges in elucidating structure and mechanism of proton pumping NADH:ubiquinone oxidoreductase (complex I). J Bioenerg Biomembr 2008. 40 (5) 475-83 Link
Morgner N, Zickermann V, Kerscher S, Wittig I, Abdrakhmanova A, Barth HD, Brutschy B, Brandt U Subunit mass fingerprinting of mitochondrial complex I. Biochim Biophys Acta 2008. 1777 (10) 1384-91 Link
Clason T, Zickermann V, Ruiz T, Brandt U, Radermacher M Direct localization of the 51 and 24 kDa subunits of mitochondrial complex I by three-dimensional difference imaging. J Struct Biol 2007. 159 (3) 433-42 Link
Kerscher S, Dröse S, Zickermann V, Brandt U The three families of respiratory NADH dehydrogenases. Results Probl Cell Differ 2008. 45 185-222 Link
Zickermann V, Zwicker K, Tocilescu MA, Kerscher S, Brandt U Characterization of a subcomplex of mitochondrial NADH:ubiquinone oxidoreductase (complex I) lacking the flavoprotein part of the N-module. Biochim Biophys Acta 2007. 1767 (5) 393-400 Link
Abdrakhmanova A, Zwicker K, Kerscher S, Zickermann V, Brandt U Tight binding of NADPH to the 39-kDa subunit of complex I is not required for catalytic activity but stabilizes the multiprotein complex. Biochim Biophys Acta 2006. 1757 (12) 1676-82 Link
Marshall D, Fisher N, Grigic L, Zickermann V, Brandt U, Shannon RJ, Hirst J, Lawrence R, Rich PR ATR-FTIR redox difference spectroscopy of Yarrowia lipolytica and bovine complex I. Biochemistry 2006. 45 (17) 5458-67 Link
Radermacher M, Ruiz T, Clason T, Benjamin S, Brandt U, Zickermann V The three-dimensional structure of complex I from Yarrowia lipolytica: a highly dynamic enzyme. J Struct Biol 2006. 154 (3) 269-79 Link
Maly T, Grgic L, Zwicker K, Zickermann V, Brandt U, Prisner T Cluster N1 of complex I from Yarrowia lipolytica studied by pulsed EPR spectroscopy. J Biol Inorg Chem 2006. 11 (3) 343-50 Link
Brandt U, Abdrakhmanova A, Zickermann V, Galkin A, Dröse S, Zwicker K, Kerscher S Structure-function relationships in mitochondrial complex I of the strictly aerobic yeast Yarrowia lipolytica. Biochem Soc Trans 2005. 33 (Pt 4) 840-4 Link
Waletko A, Zwicker K, Abdrakhmanova A, Zickermann V, Brandt U, Kerscher S Histidine 129 in the 75-kDa subunit of mitochondrial complex I from Yarrowia lipolytica is not a ligand for [Fe4S4] cluster N5 but is required for catalytic activity. J Biol Chem 2005. 280 (7) 5622-5 Link
Abdrakhmanova A, Zickermann V, Bostina M, Radermacher M, Schägger H, Kerscher S, Brandt U Subunit composition of mitochondrial complex I from the yeast Yarrowia lipolytica. Biochim Biophys Acta 2004. 1658 (1-2) 148-56 Link
Brandt U, Kerscher S, Dröse S, Zwicker K, Zickermann V Proton pumping by NADH:ubiquinone oxidoreductase. A redox driven conformational change mechanism? FEBS Lett 2003. 545 (1) 9-17 Link
Zickermann V, Bostina M, Hunte C, Ruiz T, Radermacher M, Brandt U Functional implications from an unexpected position of the 49-kDa subunit of NADH:ubiquinone oxidoreductase. J Biol Chem 2003. 278 (31) 29072-8 Link
Peng G, Fritzsch G, Zickermann V, Schägger H, Mentele R, Lottspeich F, Bostina M, Radermacher M, Huber R, Stetter KO, Michel H Isolation, characterization and electron microscopic single particle analysis of the NADH:ubiquinone oxidoreductase (complex I) from the hyperthermophilic eubacterium Aquifex aeolicus. Biochemistry 2003. 42 (10) 3032-9 Link
Kerscher S, Dröse S, Zwicker K, Zickermann V, Brandt U Yarrowia lipolytica, a yeast genetic system to study mitochondrial complex I. Biochim Biophys Acta 2002. 1555 (1-3) 83-91 Link
Kerscher S, Kashani-Poor N, Zwicker K, Zickermann V, Brandt U Exploring the catalytic core of complex I by Yarrowia lipolytica yeast genetics. J Bioenerg Biomembr 2001. 33 (3) 187-96 Link
Kashani-Poor N, Kerscher S, Zickermann V, Brandt U Efficient large scale purification of his-tagged proton translocating NADH:ubiquinone oxidoreductase (complex I) from the strictly aerobic yeast Yarrowia lipolytica. Biochim Biophys Acta 2001. 1504 (2-3) 363-70 Link
Kurki S, Zickermann V, Kervinen M, Hassinen I, Finel M Mutagenesis of three conserved Glu residues in a bacterial homologue of the ND1 subunit of complex I affects ubiquinone reduction kinetics but not inhibition by dicyclohexylcarbodiimide. Biochemistry 2000. 39 (44) 13496-502 Link
Okun JG, Zickermann V, Zwicker K, Schägger H, Brandt U Binding of detergents and inhibitors to bovine complex I - a novel purification procedure for bovine complex I retaining full inhibitor sensitivity. Biochim Biophys Acta 2000. 1459 (1) 77-87 Link
Zickermann V, Kurki S, Kervinen M, Hassinen I, Finel M The NADH oxidation domain of complex I: do bacterial and mitochondrial enzymes catalyze ferricyanide reduction similarly? Biochim Biophys Acta 2000. 1459 (1) 61-8 Link
Okun JG, Zickermann V, Brandt U Properties of the common inhibitor-binding domain in mitochondrial NADH-dehydrogenase (complex I). Biochem Soc Trans 1999. 27 (4) 596-601 Link
Björklöf K, Zickermann V, Finel M Purification of the 45 kDa, membrane bound NADH dehydrogenase of Escherichia coli (NDH-2) and analysis of its interaction with ubiquinone analogues. FEBS Lett 2000. 467 (1) 105-10 Link
Malatesta F, Nicoletti F, Zickermann V, Ludwig B, Brunori M Electron entry in a CuA mutant of cytochrome c oxidase from Paracoccus denitrificans. Conclusive evidence on the initial electron entry metal center. FEBS Lett 1998. 434 (3) 322-4 Link
Zickermann V, Barquera B, Wikström M, Finel M Analysis of the pathogenic human mitochondrial mutation ND1/3460, and mutations of strictly conserved residues in its vicinity, using the bacterium Paracoccus denitrificans. Biochemistry 1998. 37 (34) 11792-6 Link
Zickermann V, Wittershagen A, Kolbesen BO, Ludwig B Transformation of the CuA redox site in cytochrome c oxidase into a mononuclear copper center. Biochemistry 1997. 36 (11) 3232-6 Link
Zickermann V, Verkhovsky M, Morgan J, Wikström M, Anemüller S, Bill E, Steffens GC, Ludwig B Perturbation of the CuA site in cytochrome-c oxidase of Paracoccus denitrificans by replacement of Met227 with isoleucine. Eur J Biochem 1995. 234 (2) 686-93 Link
Witt H, Zickermann V, Ludwig B Site-directed mutagenesis of cytochrome c oxidase reveals two acidic residues involved in the binding of cytochrome c. Biochim Biophys Acta 1995. 1230 (1-2) 74-6 Link
With a molecular mass of almost 1MDa mitochondrial NADH:ubiquinone oxidoreductase (complex I) is the largest membrane protein complex of the respiratory chain. We have accomplished the first X-ray crystallographic analysis of the complete mitochondrial enzyme from the aerobic yeast Yarrowia lipolytica. Despite remarkable progress in structure determination of complex I we are just beginning to understand the coupling mechanism of redox-linked proton translocation. Moreover, information on the structure and function of the majority of accessory subunits is still severely limited and molecular details of complex I biogenesis are largely unknown. A number of unresolved issues calls for structural information at higher resolution and for a detailed characterization of different functional states of the enzyme complex. We combine a wide spectrum of structural and functional studies to unravel the molecular mechanism of redox-linked proton translocation by complex I.
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Figure 1: Crystallization and X-ray crystallographic analysis of mitochondrial complex I. Crystals of complex I from Y. lipolytica (A) contain all expected subunits as judged by dSDS PAGE (B). X-ray structure of mitochondrial complex I (C) |
A scaffold of accessory subunits is essential for structural integrity of mitochondrial complex I. Furthermore, accessory subunits are discussed to have a function in assembly and regulation. Mitochondrial acyl carrier proteins (ACPMs) are frequently found to be associated with complex I and have been suggested to participate in the synthesis of the essential cofactor lipoic acid. Two accessory subunits of complex I belong to the LYRM protein family. LYRM proteins have been linked with a broad range of functions predominantly in mitochondrial homeostasis. Our goal is to define the individual function of accessory complex I subunits and to explore the role of LYRM proteins in mitochondrial metabolism.
Goethe University Frankfurt
Max-von-Laue-Str. 9
60438 Frankfurt am Main
Germany
Tel: +49 (0) 69 798 29575
zickermann@med.uni-frankfurt.de
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