Washington University in St. Louis Washington University Open Scholarship Arts & Sciences Electronic Theses and Dissertations Arts & Sciences Summer 8-15-2015 Structure and Function of Cytochrome Containing Electron Transport Chain Proteins from Anoxygenic Photosynthetic Bacteria Erica Lois-Wunderlich Majumder Washington University in St. Louis Follow this and additional works at: https://openscholarship.wustl.edu/art_sci_etds Part of the Chemistry Commons Recommended Citation Majumder, Erica Lois-Wunderlich, "Structure and Function of Cytochrome Containing Electron Transport Chain Proteins from Anoxygenic Photosynthetic Bacteria" (2015). Arts & Sciences Electronic Theses and Dissertations. 567. https://openscholarship.wustl.edu/art_sci_etds/567 This Dissertation is brought to you for free and open access by the Arts & Sciences at Washington University Open Scholarship. It has been accepted for inclusion in Arts & Sciences Electronic Theses and Dissertations by an authorized administrator of Washington University Open Scholarship. For more information, please contact [email protected]. WASHINGTON UNIVERSITY IN ST. LOUIS Department of Chemistry Dissertation Examination Committee: Robert E. Blankenship, Chair Michael Gross Dewey Holten Robert Kranz Liviu Mirica Structure and Function of Cytochrome Containing Electron Transport Chain Proteins from Anoxygenic Photosynthetic Bacteria by Erica Lois-Wunderlich Majumder A dissertation presented to the Graduate School of Arts & Sciences of Washington University in partial fulfillment of the requirements for the degree of Doctor of Philosophy August 2015 St. Louis, Missouri © 2015, Erica Wunderlich Majumder Table of Contents List of Figures ................................................................................................................................. v List of Tables ................................................................................................................................ vii List of Abbreviations ................................................................................................................... viii Acknowledgments.......................................................................................................................... ix ABSTRACT OF THE DISSERTATION ....................................................................................... x Chapter 1: Diversity of Photosynthetic Cytochrome Proteins ........................................................ 1 1.1 Introduction ...................................................................................................................... 2 1.2 Cytochromes in Anoxygenic Photosynthetic Electron Transport Chains ........................ 4 1.2.1 Cytochromes in type I and II reaction centers ............................................................................. 5 Cytochromes in Type I Reaction Centers ............................................................................................. 6 Cytochromes in Type II Reaction Centers ............................................................................................ 9 1.2.2 Cytochromes in bc complexes ................................................................................................... 13 Cytochrome bc1 complex .................................................................................................................... 14 Cytochrome bc complexes .................................................................................................................. 17 Alternative Complex III ...................................................................................................................... 18 1.2.3 Soluble cytochromes in anoxygenic cyclic electron transport ................................................... 20 1.3 Conclusions ......................................................................................................................... 23 References ................................................................................................................................. 23 Chapter 2: Functional analysis and expression of the mono-heme containing cytochrome c subunit of the Alternative Complex III in Chloroflexus aurantiacus ........................................... 32 2.2 Materials and Methods ........................................................................................................ 35 2.3 Results ................................................................................................................................. 41 2.4 Discussion ........................................................................................................................... 53 References ................................................................................................................................. 58 Chapter 3: Structural analysis of diheme cytochrome c by hydrogen deuterium exchange mass spectrometry and homology modeling.......................................................................................... 63 3.1 Introduction ......................................................................................................................... 64 3.2 Materials and methods ........................................................................................................ 67 3.3 Results and Discussion ........................................................................................................ 71 3.4 Conclusions ......................................................................................................................... 88 ii References ................................................................................................................................. 89 Supplemental Figures for Chapter 3 ................................................................................................... 94 Chapter 4: Alternative Complex III from phototrophic bacteria and its electron acceptor auracyanin ..................................................................................................................................... 97 4.1 Introduction ......................................................................................................................... 98 4.2 Discovery of ACIII.............................................................................................................. 99 4.3 Protein Subunits of ACIII ................................................................................................. 100 4.3.1 ActA ......................................................................................................................................... 101 4.4 Putative Electron Transfer Mechanism for ACIII ............................................................. 114 4.5 Electron Transport Chains Involving ACIII...................................................................... 117 4.6 Auracyanins ....................................................................................................................... 119 4.7 Concluding Thoughts ........................................................................................................ 125 References ............................................................................................................................... 126 Chapter 5: Supramolecular organization of the photosynthetic complexes in the membranes of Roseiflexus castenholzii .............................................................................................................. 132 5.1 Introduction .................................................................................................................. 133 5.2 Materials and Methods ................................................................................................. 135 5.3 Results .......................................................................................................................... 139 5.4 Discussion .................................................................................................................... 150 5.5 Conclusion .................................................................................................................... 163 References ............................................................................................................................... 165 Chapter 6: Purification and Characterization of the Alternative Complex III from Roseiflexus castenholzii ................................................................................................................................. 171 6.1 Introduction .................................................................................................................. 172 6.2 Materials and Methods ................................................................................................. 173 6.2.3 Membrane Solubilization ......................................................................................................... 174 6.2.4 Protein Isolation & Purification ............................................................................................... 175 6.2.5 UV/Vis spectroscopy ............................................................................................................... 176 6.2.6 Gel Analysis ............................................................................................................................. 176 6.2.7 Mass Spectrometry ..................................................................................................................