Iron-Sulfur Clusters

Silke Leimkühler, PhD, is a Full Professor in Molecular Enzymology at the University of Potsdam in Germany. Her research is focused on molybdenum cofactor biosynthesis, molybdoenzyme enzymology, cellular sulfur transfer mechanisms for sulfur-containing biomolecule synthesis, and related subjects. Günter Schwarz is Professor for Biochemistry at the University of Cologne. His research is focused on molecular biology, protein biochemistry, enzymology, structural biology, cell biology, and more. Oliver Lenz is the Head of the Research Group Biochemistry of Gas-Converting Biocatalysts at the Technische Universität Berlin. His research is focused on the role of molecular hydrogen in the metabolism of microorganisms. Oliver Einsle is a Full Professor of Biochemistry in Freiburg, Germany and the Director of the Institute of Biochemistry in the Faculty of Chemistry and Pharmacy. His research is focused on the structural and functional characterization of metalloproteins, like nitrogenase and nitrous oxide reductase.

• Volume 1Preface xv1 ISC-based Fe–S Protein Biogenesis in Bacteria 1Béatrice Py and Frédéric Barras2 Unraveling the Complexity of the Suf-Based Fe–SBiogenesis 25Ingie Elchennawi, Claire E. Fisher, Franklin Wayne Outten, and SandrineOllagnier de Choudens3 Biogenesis of Mitochondrial Iron–Sulfur Proteins 57Ulrich Mühlenhoff, Oliver Stehling, and Roland Lill4 Iron–Sulfur Protein Maturation in the Cytosol ofEukaryotes 87Joseph J. Braymer and Antonio J. Pierik5 Mammalian Aldehyde Oxidase 135Maria Joao Romao, Guilherme Vilela-Alves, and Cristiano Mota6 Biological Formation of Sulfide 159Marion Jespersen and Tristan Wagner7 The Molybdenum Cofactor, Its Biosynthesis, and Relation toFe–S Clusters in Bacteria 193Paolo Olivieri and Silke Leimkühler8 Molybdenum Cofactor Biosynthesis in Eukaryotes 227Guenter Schwarz, Lukas Flohr, Emanuel Bruckisch, and KatrinFischer-Schrader9 NifUS Is a Two-component Toolbox Involved in AssemblingFe–S Clusters Associated with Nitrogen Fixation andBeyond 257Julia S. Martin del Campo, Shervin Shaybani, Dennis R. Dean, and Patricia C.Dos Santos10 Alternative Substrates of Nitrogenase 287Frederik V. Schmidt and Johannes G. RebeleinVolume 2Preface xvii11 Electron-Bifurcating Hydrogenases 317Gregory E. Vansuch, Effie C. Kisgeropoulos, Jonathan R. Humphreys, CarolynE. Lubner, David W. Mulder, and Paul W. KingChapter Goals 317Main Chapter Points 31711.1 Introduction 31711.1.1 Hydrogenases: From “Simple” Prototypical Architectures to Not SoSimple Electron-Bifurcating Architectures 31711.1.2 Electron Bifurcation: A Brief Overview 32011.2 Physiology, Subunit Compositions, and Reactivity 32111.2.1 Overview of Physiological Roles: Three Thematic Examples 32111.2.2 Subunit Compositions and Reactivities 32411.3 Structural and Biophysical Characterizations 33711.3.1 Structures and Cofactors of Hyd-type Electron-BifurcatingHydrogenase 33811.3.1.1 HydA and HydSL 33911.3.1.2 HydB 34011.3.1.3 HydC 34111.3.2 Spectroscopy of HydABC and HndABCD 34211.3.2.1 EPR Spectroscopy of the Fe–S Clusters in Tm HydABC 34211.3.2.2 EPR Spectroscopy of HndABCD from S. fructosivorans 34411.3.2.3 EPR and IR Spectroscopy of the H-Cluster in [FeFe]–HydABC 34411.3.3 Structures and Cofactors of MvhAGD-HdrABC 34611.3.3.1 Selenocysteine Considerations 34811.3.4 Spectroscopy of MvhAGD 34911.4 Mechanistic Proposals 34911.4.1 Electron Bifurcation: General Mechanistic Considerations 35011.4.2 [FeFe]– and [NiFe]–-HydABC: Electron Transfer Pathways 35211.4.3 [NiFe]–MvhAGD–HdrABC 35611.4.3.1 Electron Transfer Pathways 35611.4.3.2 CoM-S-S-CoB Binding and Reduction 35911.5 Forefronts 36011.6 Summary and Outlook 362Acknowledgments 363Author Contributions 363References 36412 Inhibition of [FeFe]-Hydrogenases by Small Molecules 381Claudia Brocks and Thomas HappeChapter Goals 38112.1 Historical Perspective of [FeFe]-Hydrogenase Research 38112.2 Characteristics of [FeFe]-Hydrogenases: How They Do What TheyDo 38412.3 Small Molecules Inhibit [FeFe]-Hydrogenases 38912.3.1 Diffusion of Small Molecules to the H-cluster 39012.3.2 Irreversible Attack of O2 Molecules 39212.3.3 Inhibitor Molecules that Protect [FeFe]-Hydrogenases Against O2 39412.3.3.1 CO Protects Against the O2-Initiated H-Cluster Degradation 39412.3.3.2 Formaldehyde Attacks Catalytically Important Sites of[FeFe]-Hydrogenases 39412.3.3.3 The Role of Sulfide in [FeFe]-Hydrogenases 39612.3.3.4 Cba5H – A [FeFe]-Hydrogenase that Protects Itself by Its OwnSulfide 39612.3.4 Influences of Small Molecules on the Proton Transfer Pathway 39812.4 Summary and Outlook 399References 39913 Biosynthesis of [NiFe]-Hydrogenase 407Oliver Lenz and Giorgio CasertaChapter Goals and Main Chapter Points 40713.1 [NiFe]-Hydrogenases and Their Function 40713.2 The Inorganic Catalytic Center in the [NiFe]-Hydrogenase BasicModule 40913.3 Genetic Basis of [NiFe]-Hydrogenase Maturation 40913.4 Cyanide Synthesis by HypE and HypF 41213.5 Assembly of the Fe(CN)2CO Synthon on the HypCD Complex 41413.6 Origin and Synthesis of the Carbon Monoxide Ligand 41713.7 Transfer of the Fe(CN)2CO Synthon from the HypCD Complex toApo-Hydrogenase 41913.8 Mobilization and Insertion of Nickel by HypAB 42113.9 Role of the C-Terminal Extension of the Premature Large Subunit 42513.10 Hydrogenase Maturation Intermediates Isolated from Living Cells 42713.11 Iron–Sulfur Cluster Insertion into the Small Subunit 42913.12 Subunit Oligomerization and Transport Across the CytoplasmicMembrane 43113.13 Special Case: Maturation of O2-Tolerant Membrane-Bound[NiFe]-Hydrogenases 43313.14 Conclusions 435Acknowledgments 436References 43614 [Fe]-Hydrogenase and the FeGP Cofactor Involved in theCO2-Reducing Hydrogenotrophic Methanogenic Pathway 447Seigo Shima, Joao Pedro Fernandes-Queiroz, and Masanori KanekoChapter Goals 447Main Chapter Points 44714.1 Introduction 44714.2 Unique Coenzymes Found in Methanogens 44814.3 Enzymes Involved in the Hydrogenotrophic MethanogenicPathway 44914.4 [NiFe]-Hydrogenases Involved in the HydrogenotrophicMethanogenesis 45214.5 Function of [Fe]-Hydrogenase (Hmd) 45314.6 Reconstitution of the Hmd Holoenzyme with the ExtractedCofactor 45414.7 Light Sensitivity of Hmd and Finding of the Functional Iron in theCofactor 45614.8 Structure and Properties of the FeGP Cofactor 45714.9 Photolysis Mechanism 45914.10 Crystal Structures of Hmd Apoenzymes 45914.10.1 Crystal Structure of Reconstituted Hmd Holoenzymes 46014.10.2 Crystal Structure of Reconstituted Hmd with the Substrate 46014.10.3 The Other Crystal Structures of Hmd and its Homologs 46114.11 ESI-MS Analysis for Detection of the CO/Acyl Ligands 46214.12 Biosynthesis of the FeGP Cofactor 46314.12.1 Stable Isotope Labeling Experiments 46314.12.2 The hcg Gene Cluster, Sequence Similarities, and MutationalAnalysis 46514.12.3 Structure to Function Analysis of Hcg Proteins 46614.12.3.1 HcgA 46714.12.3.2 HcgB 46714.12.3.3 HcgC 46814.12.3.4 HcgD 47014.12.3.5 HcgE 47114.12.3.6 HcgF 47214.12.3.7 HcgG 47314.12.4 In Vitro Biosynthesis 47314.12.4.1 Design of the In Vitro Biosynthesis Assay 47414.12.4.2 Confirmation of the Precursors by In Vitro Biosynthesis 47514.12.4.3 Basis of the In Vitro Complementation Assay 47614.12.4.4 In Vitro Complementation of HcgA 47614.12.4.5 In Vitro Complementation of HcgG 47714.12.5 Proposed Biosynthesis Sequence of the FeGP Cofactor 47914.13 Conclusion 479Acknowledgment 480References 48015 Catalysis by Hydrogenase 489Seigo Shima, James A. Birrell, Sven T. Stripp, Giorgio Caserta, and Oliver LenzChapter Goals and Main Chapter Points 48915.1 Catalytic Cycle of [Fe]-Hydrogenase 49015.1.1 Electronic Properties of the Iron Site of the FeGP Cofactor 49015.1.2 Enzyme Reaction Kinetics 49115.1.3 Hmd Inhibitors and Their Contribution to the CatalyticMechanism 49215.1.3.1 CO and CN− 49215.1.3.2 Isocyanides 49215.1.3.3 Cu 49315.1.3.4 Fe 49415.1.3.5 H2O2 49415.1.3.6 O2 is Reduced to H2O2 by the Hmd Reaction 49515.1.4 Semisynthetic Hmd 49615.1.4.1 Importance of the 2-OH Group of the Pyridinol Ring 49815.1.4.2 Semisynthetic [Mn]-Hydrogenase 49815.1.5 Circular Dichroism Spectroscopy 49915.1.6 Crystal Structure of Hmd with the Substrate 49915.1.7 Proposed Catalytic Mechanism of Hmd 50015.1.8 Conclusion 50215.2 Toward the Catalytic Mechanism of [FeFe]-Hydrogenase 50215.2.1 The Active Site Cofactor Is an Iron–Sulfur Cluster 50315.2.2 The Electronic Structure of the H-cluster 50415.2.3 Current State of the Catalytic Mechanism of [FeFe]-hydrogenase 50715.2.4 Toward a Consensus Catalytic Mechanism 51115.2.5 Conclusion 51215.3 Catalytic Cycle of [NiFe]-Hydrogenase 51215.3.1 Classification of [NiFe]-Hydrogenases: The Complex Trait of O2Tolerance 51415.3.2 [NiFe]-Hydrogenase Active Site States 51615.3.2.1 CO-Bound States 51715.3.2.2 Hydroxy-Bridged States 51715.3.2.3 Unusual [NiFe] Site Arrangements 51915.3.3 Catalytic Intermediates 52015.3.3.1 Nia-S 52015.3.3.2 Nia-SR 52215.3.3.3 Nia-C 52315.3.3.4 Nia-L 52415.3.4 Conclusions 525Acknowledgment 525References 52616 Macromolecular Crystallography 541Konstantin Bikbaev and Ingrid SpanChapter Goals 541Main Chapter Points 54116.1 Introduction 54216.2 Crystallization of Macromolecules 54416.3 Symmetry and the Unit Cell 54816.4 Diffraction and Interpretation of Diffraction Patterns 55016.5 Data Collection and Processing 55216.6 X-ray structure analysis 55416.6.1 Isomorphous Replacement 55516.6.2 Anomalous Diffraction 55616.6.3 Molecular Replacement 55716.6.4 Structure Refinement 55716.7 Time-Resolved Crystallography 56216.8 Nuances of Fe–S Protein Crystallography 56516.9 Conclusions 566References 56617 Vibrational and Mössbauer Spectroscopic Techniques to StudyIron–Sulfur Clusters 571Christian Lorent, Giorgio Caserta, Volker Schünemann, and Ingo ZebgerChapter Goals 571Main Chapter Points 57117.1 Vibrational Spectroscopy 57217.1.1 Normal Modes 57317.1.2 Vibrational Spectroscopic Techniques 57317.2 Infrared Spectroscopy 57417.2.1 MIR Spectroscopy 57517.2.2 FIR Spectroscopy 57817.2.3 Conclusion 57917.3 Raman Spectroscopy 57917.3.1 Resonance Raman Spectroscopy 58117.3.2 Studying [Fe–S] Clusters by Resonance Raman Spectroscopy 58217.3.3 Rubredoxin 58217.3.4 [2Fe–2S] Clusters 58317.3.5 [3Fe–4S] Clusters 58517.3.6 [4Fe–4S] Clusters 58617.3.7 Studying Hydrogenases by Resonance Raman Spectroscopy 58717.3.8 Conclusion 58817.4 Mössbauer-Based Spectroscopic Techniques 58817.4.1 Mössbauer Spectroscopy 58817.4.2 Mössbauer Spectroscopy from Single Fe(S−Cys)4 Sites up to [4Fe–4S]Clusters 59217.4.3 Complex Iron–Sulfur Clusters: Hydrogenases and Nitrogenases 59517.4.4 Conclusion 59617.5 Nuclear Resonance Vibrational Spectroscopy 59617.5.1 Theoretical Background 59717.5.2 [Fe(S–Cys)4] Metal Site: The Case Study of Rubredoxin 59917.5.3 [2Fe–2S] Clusters: 4xCys, 2xCys–2xHis, and 3xCys–1xHis Ligations 59917.5.4 [3Fe–4S] Clusters 60117.5.5 [4Fe–4S] Clusters 60217.5.6 Binding of NO to [Fe–S] Clusters 60217.5.7 The Nitrogenase FeMe Cofactor 60317.5.8 [NiFe]- and [FeFe]-Hydrogenases 60517.5.9 Conclusion 606References 60618 The Multisite Microstate Model: A Theoretical Approach forAnalyzing Charge Transfer in Proteins 617G. Matthias Ullmann and Rajeev Ranjan RoyMain Chapter Points 61718.1 Introduction 61718.2 Binding of Ligands to a Receptor 62018.3 Analyzing Biomolecular Systems 62218.3.1 General Considerations 62218.3.2 Thermodynamic Analysis 62418.3.3 Kinetic Analysis 62518.4 Modeling Protein Electrostatics Using the Poisson–BoltzmannEquation 62618.4.1 Conceptual Model 62618.4.2 The Mathematical Model 62718.4.3 Electrostatic Potentials and Electrostatic Energies 62918.4.4 Defining the Low-Dielectric Cavity of the Protein 63118.4.5 Fitting Quantum Chemical Electrostatic Potentials to PointCharges 63318.4.6 Problems and Shortcomings of the Poisson–Boltzmann Model 63518.5 Electrostatic Calculations of the Energy Parameter 63618.6 Practical Hints to Construct a Multisite System 63918.6.1 Molecular Structures as a Basis of Detailed Calculations 63918.6.2 How to Choose Sites, Model Compounds, and Model CompoundEnergies 64018.7 Conclusions 642Acknowledgement 643References 64319 Structural and Functional Bioinorganic Model Chemistry ofFe–S Clusters – Synthesis and Analysis 649Benedict Josua Elvers and Carola SchulzkeChapter Goals and Main Chapter Points 64919.1 Introduction 64919.2 Fe–S Cluster Motifs: Natural Occurrence, Reactivity, and ArtificialSynthesis 65219.3 [FeS4] 65319.3.1 Biological Relevance 65319.3.2 Synthetic Model Complexes 65419.4 [Fe2S2] Clusters 65719.4.1 Biological Relevance of [Fe2S2] Clusters 65819.4.2 Synthetic Model Complexes of [Fe2S2] Clusters 65819.5 [Fe4S4] and [Fe3S4] Clusters 66519.5.1 Biological Relevance of [Fe4S4] and [Fe3S4] Clusters 66519.5.2 Synthetic Model Complexes of [Fe4S4] 66719.5.3 Synthetic Model Complexes of [Fe3S4] 67619.6 Clusters of Higher Nuclearity 67919.6.1 Hydrogenase and Nitrogenase 68019.6.2 Model Chemistry for Clusters of Higher Nuclearity 68319.7 Conclusion 691References 69220 Metal-Dependent Formate Dehydrogenases and TheirInterplay and Relationship to Iron–Sulfur Clusters 705Benjamin R. DuffusChapter Goals 705Main Chapter Points 70520.1 Formate: General Considerations 70620.2 Metal-Independent FDHs 70720.3 Metal-Dependent FDHs 70820.4 Metal-Dependent Formate Dehydrogenases: Common Modular CatalyticUnit 71120.5 FDH and Link of bis-MGD to a [4Fe–4S] Cluster 71220.6 Link of the FDH bis-MGD Cofactor with Extended Fe-S Cluster ElectronChains 71420.7 FDH that Employ Ferredoxin as Electron Acceptor 71520.8 E. coli Metal-Dependent Formate Dehydrogenases 71520.9 Formate Hydrogenlyase Complex 71620.10 NAD+-Reducing, Metal-Dependent FDHs 71720.11 W-Containing FDHs from Sulfate-Reducing Bacteria 71920.12 FDH: D. vulgaris Hildenborough 71920.13 Tungsten vs. Molybdenum FDH 72020.14 Metal-Dependent Formate Dehydrogenase from Moorella thermoacetica:Variance and Codependence of Metal Ions 72120.15 Formylmethanofuran Dehydrogenases 72120.16 FDHs in Methanogens 72220.17 FDH O2 Sensitivity: Inhibition and Enzymatic Activation 72420.18 FDH and Electron Bifurcation vs. O2 Tolerance 72520.19 FDH in H2-Dependent CO2 Reduction (HDCR) 72720.20 Conclusion 728Acknowledgments 729References 729Index 747