University of Wollongong University of Wollongong Research Online Research Online University of Wollongong Thesis Collection 2017+ University of Wollongong Thesis Collections 2019 Synthesis and Quadru[.]
University of Wollongong Research Online University of Wollongong Thesis Collection 2017+ University of Wollongong Thesis Collections 2019 Synthesis and Quadruplex DNA Binding Properties of Novel Nickel Schiff Base Complexes Son Quynh Thai Pham University of Wollongong Follow this and additional works at: https://ro.uow.edu.au/theses1 University of Wollongong Copyright Warning You may print or download ONE copy of this document for the purpose of your own research or study The University does not authorise you to copy, communicate or otherwise make available electronically to any other person any copyright material contained on this site You are reminded of the following: This work is copyright Apart from any use permitted under the Copyright Act 1968, no part of this work may be reproduced by any process, nor may any other exclusive right be exercised, without the permission of the author Copyright owners are entitled to take legal action against persons who infringe their copyright A reproduction of material that is protected by copyright may be a copyright infringement A court may impose penalties and award damages in relation to offences and infringements relating to copyright material Higher penalties may apply, and higher damages may be awarded, for offences and infringements involving the conversion of material into digital or electronic form Unless otherwise indicated, the views expressed in this thesis are those of the author and not necessarily represent the views of the University of Wollongong Recommended Citation Pham, Son Quynh Thai, Synthesis and Quadruplex DNA Binding Properties of Novel Nickel Schiff Base Complexes, Doctor of Philosophy thesis, School of Chemistry and Molecular Bioscience, University of Wollongong, 2019 https://ro.uow.edu.au/theses1/639 Research Online is the open access institutional repository for the University of Wollongong For further information contact the UOW Library: research-pubs@uow.edu.au Synthesis and Quadruplex DNA Binding Properties of Novel Nickel Schiff Base Complexes Son Quynh Thai Pham Master of Science (Chemistry) This thesis is presented as part of the requirements for the award of the Degree of Doctor of Philosophy of the University of Wollongong School of Chemistry and Molecular Bioscience March 2019 Declaration I, Son Quynh Thai Pham, declare that this thesis, submitted in partial fulfilment of the requirements for the award of Doctor of Philosophy, in the School of Chemistry and Molecular Bioscience, University of Wollongong, is wholly my own work unless otherwise referenced or acknowledged This work has not been submitted for qualification at any other academic institution Son Quynh Thai Pham 29th March 2019 i Abstract Thirteen novel Schiff base complexes have been successfully synthesised through reactions of substituted benzophenones with different diamines in the presence of nickel(II) acetate These precursor complexes then were successfully alkylated using 1-(2-choroethyl)piperidine hydrochloride to form a series of novel nickel complexes bearing dimethylenepiperidine pendant groups The nickel complexes with the pendant groups were sufficiently soluble in water to enable them to be used in DNA binding experiments All new complexes were fully characterised using NMR spectroscopy, Electrospray Ionisation Mass Spectrometry (ESI-MS) and elemental microanalysis In addition, the solid state structures of eight complexes were determined using X-ray crystallography Various techniques including ESI-MS, Circular Dichroism (CD), UV-Vis spectrophotometry, Fluorescence Indicator Displacement (FID), Fluorescence Resonance Energy Transfer (FRET) melting assays and molecular docking were employed to investigate the effects of structural variations amongst the nickel Schiff base complexes on their DNA binding properties DNA binding studies were performed using the tetramolecular G-quadruplex Q4, the unimolecular Gquadruplexes Q1 and c-kit1, the fluorescently labelled unimolecular G-quadruplex F21T, and the double stranded DNA molecule D2 Experiments involving Q1 were performed after it was annealed under specific conditions to afford parallel, antiparallel and hybrid topologies The results of DNA binding studies indicated that varying the number of pendant groups appended to the Schiff base scaffold resulted in the largest changes to DNA affinity and selectivity For example, complex (89), carrying four pendant groups exhibited strong affinities towards many kinds of Gii quadruplex DNA, including parallel Q4 and Q1, anti-parallel Q1, c-kit1 and antiparallel F21T DNA binding studies performed using five isomeric nickel Schiff base complexes containing two pendant groups in different locations also showed significant variations in the strength of interactions with some G-quadruplexes, such as parallel Q4, parallel c-kit1, and anti-parallel F21T Modifying the diamine moieties in the top half of the nickel Schiff base complexes, and introducing asymmetry into their structures, resulted generally in smaller changes to DNA affinities and selectivities iii Acknowledgement First and foremost, I would like to send the greatest thanks to my supervisor Prof Stephen Ralph for your continuous guidance and support before and during my PhD project You are my role model of hard working, enthusiasm and dedication Doing research is never an easy job, particularly when the experiments not go well However, it seems you always have the answers for all the problems Most importantly, you always make me feel better about myself and about my works which makes doing research is less stressful and more enjoyable Thank you for opening the door and letting me into the world of science and for everything you have done for me I also would like to thank my co-supervisor Dr Celine Kelso who always put me on the top of her job list Thank you very much for walking me through every steps and sharing your expertise in the field of mass spectrometry My special thanks go to Prof Jenny Beck for your generosity, silent supports, and willingness to help me not only in my research project but also any other problems that I have encountered during the time I have been in Australia Also, I am particularly grateful to Dr Kimberley Davis for her training, detailed instructions as well as valuable advice To Dr Christopher Richardson and Dr Anthony C Willis, thank you very much for your help with X-ray crystallography experiments To Assoc Prof Haibo Yu, Dr Nguyen Thuy Viet Phuong and the computational chemistry group at UoW, thank you very much for your help with my modelling experiments In addition, thanks to Dr Wilford Lie for your training with NMR techniques and thanks Dr Monica Birrento for iv your assistance with performing FRET experiments Thank you to the technical and administrative staffs at the School of Chemistry and Molecular Bioscience of UoW for your excellent support during my PhD project Special thanks are extended to the University of Wollongong for providing a full scholarship for my PhD degree Moreover, an important acknowledgement will go to Australian taxpayers for offering me the full scholarship for my master degree which was a very important step for me to continue my doctoral study at UoW I also would like to thank my wonderful relatives and friends in Australia and at home: Aunt Lieu, Aunt Hai, Uncle Son, Uncle Minh, Uncle Ba, Tram, Kevin, Chip, Luong, Chau, Trang, Nghia, Han, Thi, Hong, Paul, Jessica and Harun Thanks Aunt Lieu, Aunt Hai, Uncle Son and Tram for your support from Vietnam and always being willing to help Thanks Kevin and Chip for dragging me out my room and showing me around New South Wales Thank you to Harun and Paul for the fishing trips, sharing delicious foods and your interesting cultures Finally, thanks to my parent for having me Most importantly, thanks Mom for all of your sacrifices since the day Dad passed away thirty years ago Thank you to my sister and my brother in law for your continuous support and for taking care of Mom when I have been away from home v Table of Contents Chapter : Introduction 1.1 DNA replication and cancer 1.2 Telomeres, telomerase and their role in cancer cell growth 1.3 Telomeres and telomerase as potential therapeutic targets for cancer treatment 1.4 Quadruplex DNA 15 1.5 An overview of G-quadruplex DNA binding agent 18 1.5.1 Organic qDNA binding agents 20 1.5.2 Metal complexes as G-quadruplex binding agents 22 1.5.3 Metal Schiff base complexes 35 1.6 Methods for investigating the G-quadruplex DNA binding properties of metal complexes 41 1.6.1 Electrospray Ionisation Mass Spectrometry (ESI-MS) 41 1.6.2 Circular Dichroism (CD) spectroscopy 44 1.6.3 Absorption spectrophotometry 47 1.6.4 Fluorescence Intercalator Displacement (FID) assays 49 1.6.5 Fluorescence Resonance Energy Transfer (FRET) 50 1.6.6 Molecular docking 52 1.7 Thesis objectives 55 Chapter : Materials and methods 60 2.1 Materials 60 2.1.1 Reagents used for synthesis 60 2.1.2 Reagents used for analytical techniques 60 2.2 Characterisation of metal complexes 61 2.2.1 General characterisation 61 2.2.2 Crystallographic characterisation 62 2.3 Preparation of solutions of metal complexes and oligonucleotides for DNAbinding studies 64 2.3.1 Preparation of metal complex stock solutions 64 2.3.2 Purification of oligonucleotides 65 2.3.3 Preparation of double stranded DNA and quadruplex DNA 66 2.4 Mass spectrometry DNA–binding experiments 67 2.5 Circular dichroism (CD) DNA-binding experiments 69 2.6 Absorption spectrophotometry DNA-binding experiments 70 2.6.1 Absorption titrations 70 2.6.2 DNA melting experiments 71 2.7 Fluorescence intercalator displacement (FID) assays 71 2.8 Fluorescence resonance energy transfer (FRET) DNA-binding assays 73 2.9 Molecular docking experiments 74 vi Chapter : Synthesis and structural characterisation of nickel complexes of benzophenone Schiff base ligands 78 3.1 Introduction 78 3.2 Overview of synthetic procedures 79 3.2.1 Synthetic reactions 79 3.2.2 Purification procedures 81 3.3 Results and discussion 81 3.3.1 Synthesis of nickel complexes containing different diamine groups 81 3.3.2 Synthesis of isomeric nickel complexes 99 3.3.3 Synthesis of nickel complexes with different numbers of pendant groups 115 3.3.4 Synthesis of nickel complexes with asymmetric structures 122 3.4 X-ray crystallographic characterisation of nickel complexes 133 3.4.1 Solid-state structures of non-alkylated nickel complexes 133 3.4.2 Solid-state structures of alkylated nickel complexes 140 Chapter : Effect of varying the number of pendant groups on DNA binding properties 146 4.1 Introduction and scope 146 4.2 Results and discussion 148 4.2.1 DNA binding studies performed using ESI mass spectrometry 148 4.2.2 DNA binding studies performed using CD spectroscopy 152 4.2.3 DNA binding studies performed using UV-Vis spectroscopy 169 4.2.4 DNA binding studies performed using FRET melting assays 175 4.2.5 DNA binding studies performed using FID assays 178 4.2.6 DNA binding studies performed using molecular docking 180 4.3 Summary 186 Chapter : Effect of varying the positions of pendant groups on DNA binding properties 189 5.1 Introduction and scope 189 5.2 Results and discussion 191 5.2.1 DNA binding studies performed using ESI mass spectrometry 191 5.2.2 DNA binding studies using CD spectroscopy 195 5.2.3 DNA binding studies performed using UV-Vis spectroscopy 208 5.2.4 DNA binding studies performed using FRET melting assays 211 5.2.5 DNA binding studies performed using FID assays 212 5.2.6 DNA binding studies performed using molecular docking 215 5.3 Summary 218 Chapter : Effect of varying the diamine moiety on DNA binding properties 220 6.1 Introduction and scope 220 6.2 Results and discussion 221 6.2.1 DNA binding studies performed using ESI mass spectrometry 221 6.2.2 DNA binding studies performed using CD spectroscopy 225 6.2.3 DNA binding studies performed using UV-Vis spectrophotometry 235 vii 6.2.4 DNA binding studies performed using FRET melting assays 238 6.2.5 DNA binding studies performed using FID assays 240 6.2.6 DNA binding studies performed using molecular docking 242 6.3 Summary 245 Chapter : Effect of introducing asymmetry on DNA binding properties 246 7.1 Introduction and scope 246 7.2 Results and discussion 247 7.2.1 DNA binding studies performed using ESI mass spectrometry 247 7.2.2 DNA binding studies performed using CD spectroscopy 251 7.2.3 DNA binding studies performed using UV-Vis spectrophotometry 263 7.2.4 DNA binding studies performed using FRET melting assays 266 7.2.5 DNA binding studies performed using FID assays 268 7.2.6 DNA binding studies performed using molecular docking 269 7.3 Summary 271 Chapter : Conclusions and future directions 273 8.1 Conclusions 273 8.2 Future directions 278 Supplementary Figures 281 References 303 viii ... Effect of addition of nickel Schiff base complexes on the CD spectrum of ckit1 233 Table 6.6 Effect of addition of nickel Schiff base complexes on the CD spectrum of dsDNA D2 ... Effect of addition of nickel Schiff base complexes with different numbers of pendant groups on the CD spectrum of c-kit1 164 xv Table 4.6 Effect of addition of nickel Schiff base complexes. . .Synthesis and Quadruplex DNA Binding Properties of Novel Nickel Schiff Base Complexes Son Quynh Thai Pham Master of Science (Chemistry) This thesis is presented as part of the requirements