Georgia State University ScholarWorks @ Georgia State University Chemistry Dissertations Department of Chemistry Fall 9-23-2013 DNA Interactions and Photocleavage by Anthracene, Acridine, and Carbocyanine-Based Chromophores Carla Mapp Follow this and additional works at: https://scholarworks.gsu.edu/chemistry_diss Recommended Citation Mapp, Carla, "DNA Interactions and Photocleavage by Anthracene, Acridine, and Carbocyanine-Based Chromophores." Dissertation, Georgia State University, 2013. https://scholarworks.gsu.edu/chemistry_diss/82 This Dissertation is brought to you for free and open access by the Department of Chemistry at ScholarWorks @ Georgia State University. It has been accepted for inclusion in Chemistry Dissertations by an authorized administrator of ScholarWorks @ Georgia State University. For more information, please contact [email protected]. DNA INTERACTIONS AND PHOTOCLEAVAGE BY ANTHRACENE, ACRIDINE, AND CARBOCYANINE-BASED CHROMOPHORES by CARLA TERRY MAPP Under the Direction of Dr. Kathryn B. Grant ABSTRACT The interaction of small molecules with DNA has been extensively studied and has produced a large catalogue of molecules that non-covalently bind to DNA though groove binding, intercalation, electrostatics, or a combination of these binding modes. Anthracene, acridine, and carbocyanine-based chromophores have been examined for their DNA binding properties and photo-reactivities. Their planar aromatic structures make them ideal chromophores that can be used to probe DNA structural interactions and binding patterns. We have studied DNA binding and photocleavgage properties of a bisacridine chromophore joined by a 2,6-bis(aminomethyl)pyridine copper-binding linker (Chapter II), a series of 9-aminomethyl anthracene chromophores (Chapters III and IV), both under conditions of high and low ionic strength, as well as a series of pentamethine linked symmetrical carbocyanine dyes (Chapter V). In Chapter II we present data showing that high ionic strength efficiently increases copper(II)- dependent photocleavage of plasmid DNA by the bisacridine based chromophore (419 nm, pH 7.0). In Chapters III and IV, using an pyridine N-substituted 9-(aminomethyl)anthracene (Chapter III), a bis-9-(aminomethyl)anthracene, and its mono 9-(aminomethyl)anthracene analogue (Chapter IV), pUC19 plasmid DNA was photo-converted to highly diffuse DNA fragments (350 nm, pH 7.0) in the presence of 150 mM NaCl and 260 mM KCl. Spectroscopic analyses suggest that the combination of salts promotes a change in DNA helical structure that initiate a switch in anthracene binding mode from intercalation to an external or groove binding interactions. The alteration in DNA structure and binding mode leads to an increase in the anthracene-sensitized production of DNA damaging reactive oxygen species. Finally, in Chapter V, pUC19 plasmid DNA is converted to its nicked circular and linear forms following irradiation of a series of pentamethine linked symmetrical carbocyanines (red light, pH 7.0). The data suggest that the relative levels of photocleavage arise from the different substituents on the nitrogen alkyl side chain and the pentamethine linker. INDEX WORDS: Anthracene, Photocleavage, Acridine, Intercalation, Groove binding, Photodynamic therapy DNA PHOTCLEAVAGE AND INTERACTIONS BY ANTHRACENE, ACRIDINE, AND CARBOCYANINE BASED CHROMOPHORES by CARLA TERRY MAPP A Dissertation Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in the College of Arts and Sciences Georgia State University 2013 Copyright by Carla Mapp 2013 DNA PHOTCLEAVAGE AND INTERACTIONS BY ANTHRACENE, ACRIDINE, AND CARBOCYANINE BASED CHROMOPHORES by CARLA TERRY MAPP Committee Chair: Dr. Kathryn B. Grant Committee: Dr. David Wilson Dr. Donald Hamelberg Electronic Version Approved: Office of Graduate Studies College of Arts and Sciences Georgia State University December 2013 iv DEDICATION I dedicate this dissertation to my daughter Madison, my little nugget. Always believe in yourself, reach for the stars, and you will achieve your dreams. v ACKNOWLEDGEMENTS This dissertation would not have been possible without the guidance and the help of several individuals who in one way or another contributed and extended their valuable assistance in the preparation and completion of this study. To my lab mates, both past and present, it was a pleasure to work with you all. To Orapin, thank you for contributing your hard work and insight for Chapter II. I will never forget our wonderful foodie adventures. To Dominique and Blessing I count myself lucky to to call you colleagues. And I applaud you girls for increasing diversity in the biochemistry industry. To my parents and brothers, thank you for the love, support and understanding that you have given during the long years of my education. To my wonderful, generous and very supportive husband, you have been here with me from the start of this journey and somehow kept me sane through it all. Thank you. To my invaluable network of supportive and loving friends you guys have been absolutely fabulous through this entire journey. To my committee chair and advisor Dr. Kathy Grant, Thank you for all the years of advice and guidance that you have given me. I count myself very lucky to have worked with you. To my committee members, Dr. Dixon, Dr. Hamelberg, and Dr. Wilson, for their encouraging words, thoughtful criticism, time and attention during many busy semesters. Dr. Dixon, the advice, both career and research related, that you have given over the years has been invaluable to me. I offer my sincerest thanks to you. To Dr. Wilson, thank you for allowing me to utilize your laboratory space and equipment and most of all your troubleshooting insight. vi TABLE OF CONTENTS ACKNOWLEDGEMENTS v LIST OF TABLES xii LIST OF FIGURES xiii LIST OF SCHEMES xviii CHAPTER 1. DNA STRUCTURE AND FUNCTION 1 1.1. The Double Helix: DNA Sturcutre 1 1.2. Non-Covalent DNA Molecular Interactions 3 1.2.1. Electrostatic Interactions 3 1.2.2. Groove Binding 3 1.2.3. Intercalation 6 1.3. Photodynamic Therapy and DNA Photocleavage 6 1.4. Nucleic Acid Photocleavage 9 1.4.1. Strand Scission due to Oxidative Damage of the Nucleobases 9 1.4.2. Strand Scission due to Oxidative Damage of Deoxyribose 11 1.5. DNA Interactions with Anthracene, Acridine, and Carbocyanine Chromophores 13 1.6. Specific Aims 17 1.7. Dissertation Summary 18 1.8. References 22 vii CHAPTER 2. SYNTHESIS AND DNA PHOTOCLEAVAGE BY A PYRIDINE-LINKED BIS-ACRIDINE CHROMOPHORE IN THE PRESENCE OF COPPER(II): IONIC STRENGTH EFFECTS. 27 2.1. Abstract 27 2.2. Introduction 27 2.3. Results and Discussion 29 2.3.1. Synthesis 29 2.3.2. Photocleavage Experiments 30 2.3.3. Inhibition of DNA Photocleavage 32 2.3.4. Ionic Strength Effects 33 2.3.5. UV-Visible Spectral Analysis 35 2.3.6. Thermal Melting Studies 36 2.4. Conclusion 37 2.5. References 39 2.6. Supplementary Data 42 CHAPTER 3. PHYSIOLOGICALLY RELEVANT CONCENTRATIONS OF NaCl AND KCl INCREASE DNA PHOTOCLEAVAGE BY N-SUBSTITUTED 9- AMINOMETHYLANTHRACENE DYE. 48 3.1. Abstract 48 3.2. Introduction 49 3.3. Experimental Procedures 51 3.3.1. Materials and Methods 51 3.3.2. Synthesis of (6-((methyl(2-(methylamino)ethyl)amino)methyl)pyridin- viii 2-yl)methanol (2) 52 3.3.3. Synthesis of (6-(((2-((anthracen-9-ylmethyl)(methyl)amino) ethyl)(methyl)amino)methyl)pyridin-2-yl)methanol (3) 53 3.3.4. Synthesis of N-(anthracen-9-ylmethyl)-N’-((6-((dimethylamino)methyl) pyridin-2-yl)methyl)-N,N’-dimethylethane-1,2-diamine (4) 54 3.3.5. Photocleavage of Supercoiled Plasmid DNA 55 3.3.6. Circular Dichroism Analysis 56 3.3.7. UV-Visible Absoprtion Titrations 57 3.3.8. Fluorescence Measurements 57 3.3.9. Thermal Melting of DNA 57 3.3.10. Chemically Incuded Changes in DNA Photocleavage 58 3.4. Results and Discussion 59 3.4.1. Synthesis of N-(anthracen-9-ylmethyl)-N’-((6((dimethylamino)methyl )pyridin-2-yl)methyl)-N,N’-dimethylethane-1,2-diamine (4) 59 3.4.2. Photocleavage of Supercoiled Plasmid DNA 60 3.4.3. Circular Dichroism Analysis 66 3.4.4. Absoprtion Titrations and ICD Data 69 3.4.5. Fluorescence Measurements 75 3.4.6. Thermal Denaturation Experiments 78 3.4.7. Chemically Incuded Changes in DNA Photocleavage 82 3.5. Summary and General Discussion 84 3.6. Conclusion 87 ix 3.7. Supporting Information 88 3.7.1. Discussion 90 3.8. References 97 3.9. Supporting Information References 105 CHAPTER 4. BIS- AND MONO-9-AMINOMETHYLANTHRACENE DYES: THE EFFECTS OF CHLORIDE SALTS ON DNA INTERACTIONS AND PHOTOCLEAVAGE. 107 4.1. Abstract 107 4.2. Introduction 108 4.3. Results and Discussion 110 4.3.1. Preparation of Anthracene Dyes 2 and 4 110 4.3.2. DNA Photocleavage 111 4.3.3. UV-visible Absorption Spectroscopy 113 4.3.4. Viscometric Titration 117 4.3.5. Circular Dichroism 120 4.3.6. Thermal Melting Titrations 123 4.3.7. Summary and General Discussion 126 4.3.8. Concluding Remarks 130 4.4. Experimental Section 131 4.4.1. General 131 4.4.2. Synthesis of 1,10-bis(9-anthracenemethyl)-1,4,7,10-tetraazadecane tetrahydrochloride (2) 131 x 4.4.3. Synthesis of N-(9-anthracenemethyl) ethylenediamine dihydrochloride (4) 132 4.4.4. DNA Photocleavage 133 4.4.5. UV-visible Absorption Spectrophotometry 134 4.4.6. Viscometric Titrations 134 4.4.7. Circular Dichroism 135 4.4.8. Thermal Melting Titration 135 4.5. Acknowledgements 135 4.6. Appendix A. Supplementary Data 136 4.7. References 142 CHAPTER 5. OXIDATIVE CLEAVAGE OF DNA BY PENTAMETHINE CYANINE DYES WITH LONG-WAVELENGTH VISIBLE LIGHT. 150 5.1. Abstract 150 5.2. Introduction 150 5.3. Results and Discussion 153 5.3.1. UV-visible Absorbance and Fluorescence Analysis 153 5.3.2. Photocleavage of Supercoiled Plasmid DNA at 575 nm, 588 nm, 623 nm, and 700 nm 156 5.3.3. Inhibition of DNA Photocleavage 159 5.3.4.