Synthesis and Quadruplex DNA Binding Properties of Novel Nickel Schiff Base Complexes :Doctor of Philosophy - Major: Chemistry and Molecular bioscience

University of Wollongong

Research Online

University of Wollongong Thesis Collection

2017+ University of Wollongong Thesis Collections

2019

Synthesis and Quadruplex DN Synthesis and Quadruplex DNA Binding Pr A Binding Properties of No ties of Novel Nick el Nickel Schiff el 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 do 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: [email protected]

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

i

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

ii

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 G￾quadruplexes 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, anti￾parallel 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 G-

iii

quadruplex DNA, including parallel Q4 and Q1, anti-parallel Q1, c-kit1 and anti￾parallel 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.

iv

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 do 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

v

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.

vi

Table of Contents

Chapter 1 : Introduction........................................................................................... 1

1.1 DNA replication and cancer.............................................................................. 1

1.2 Telomeres, telomerase and their role in cancer cell growth.............................. 3

1.3 Telomeres and telomerase as potential therapeutic targets for cancer

treatment ................................................................................................................ 7

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 2 : 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 DNA￾binding 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

vii

Chapter 3 : 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 4 : 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 5 : 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 6 : 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