This thesis is concerned with the electrochemical and spectroelectrochemical characterisation of cyano (CN) and trifluoromethyl (CF3) substituted polypyridine ligands and their metal complexes. The ligands investigated were X-CN-py (X = 3, 4 and 5, py = pyridine), X,X´-(CN)2-bpy) (X,X´ = 3,3´, 4,4´ and 5,5´ bpy = 2,2´- bipyridine) and X,X´-(CF3)2-bpy (X,X´ = 3,3´, 4,4´ and 5,5´). The Pt(II) complexes of the X-CN-py and X,X´-(CN)2-bpy ligands were studied along with the Fe(II) complexes of the X,X´-(CN)2-bpy and X,X´-(CF3)2-bpy ligands. Electrochemical studies of the X-CN-py ligands indicated that 2-CN-py and 4-CN-py have one reversible 1e- reduction and 3-CN-py has one quasi-reversible reduction. 4-CN-py is more easily reduced than 2-CN-py. EPR experiments on [2-CN-py]1- and [4-CN-py]1- combined with DFT calculations have indicated that the semi-occupied molecular orbital (SOMO) is delocalised over the entire molecule. The complex [Pt(4-CN-py)2Cl2] undergoes two reversible 1e- reductions, attributed to the sequential reduction of the two 4-CN-py ligands. [Pt(3-CN-py)2Cl2] was found to exhibit one irreversible reduction. The electrochemistry of the X,X´-(CN)2-bpy ligands indicated that 3,3´-(CN)2-bpy and 5,5´-(CN)2-bpy have two reversible 1e- reductions while 4,4´-(CN)2-bpy has only one reversible 1e- reduction. The reductions of 5,5´-(CN)2-bpy occur at significantly less negative potentials than those in 3,3´ or 4,4´ analogue. In-situ UV/Vis/NIR and EPR experiments on the X,X´-(CN)2-bpy ligands indicate that the reduction electron is delocalised over both the CN-py rings with the SOMO being spread across the entire molecule. [Pt(3,3´-(CN)2-bpy)Cl2] and [Pt(5,5´-(CN)2-bpy)Cl2] undergo two reversible 1e- reductions, attributed to the sequential reduction of the X,X´-(CN)2-bpy ligand. 4,4´-(CF3)2-bpy and 5,5´-(CF3)2-bpy, whose X-ray crystallographically determined structures are reported, both have one reversible 1e- reduction while 3,3´-(CF3)2-bpy exhibits an irreversible reduction. Again the 5,5´ analogue is the most easily reduced. Spectroelectrochemical results indicate that the reduction electron enters a SOMO that is delocalised over both the CF3-py rings. Substitution in the 5,5´ positions is determined to be electronically most significant. Complexes of the general formula [Fe(II)(X2-bpy)3][BF4] (where X = CN or CF3 in the 4,4´ or 5,5´ positions) exhibit three reversible ligand based reduction processes and a metal based oxidation. UV/Vis/NIR and EPR studies have confirmed that the reduction electron in each case enters a molecular orbital that is predominantly based on a bpy ligand. The redox potentials of 4-CN-py and 5,5´-(CN)2-bpy are solvent dependent with 1e- reduction of 4-CN-py and the two 1e- reductions of 5,5´-(CN)2-bpy moving to less negative potentials as the acceptor number (AN) of the solvent increases. A computational model has been developed for the study of the electronic properties of substituted bipyridines using DFT methods. This model has been used to aid the analysis of the EPR and UV/Vis/NIR spectra of the X2-bpys studied.

The function of biomolecules is very sensitive to structure. Folding in proteins and nucleic acids is a hierarchical process progressing from primary to secondary, then tertiary, and finally, quaternary structures. RNA in its folded form performs a variety of biological activities. Obtaining intramolecular distance measurements makes it possible to generate structural models along the folding pathway that may be related to the overall function of the molecule. Distances can be measured by Site-Directed Spin-Labeling (SDSL), in which nitroxyl spin-label probes are attached and observed by EPR spectroscopy. Spin-labels can provide information concerning structure and conformational changes because they are particularly sensitive to molecular motion and interspin distances. Continuous-wave EPR spectroscopy has been commonly applied to detect and monitor nitroxide spin-label probes within biological systems. A previous published SDSL study from this laboratory investigated a 10-mer RNA duplex model system with spin-label probe succinimdyl-2,2,5,5-tetramethyl-3-pyrroline-1-oxyl-carboxylate; however, an increased spin-labeling efficiency was observed with an isocyanate derivative of tetramethylpiperidyl-N-oxy (TEMPO). In this thesis, a 4-isocyano TEMPO spin-label probe replaced the previously used succinimdyl-2,2,5,5-tetramethyl-3-pyrroline-1-oxyl-carboxylate in 10-mer SDSL studies. The influence of labeling with the 4-iscocyano TEMPO spin-label in a 10-mer RNA model system was investigated with thermal denaturation, Matrix Assisted Laser Desorption Time of Flight Mass Spectrometry (MALDI-TOF-MS), Electron Paramagnetic Resonance (EPR) spectroscopy, and reverse phase high performance liquid chromatography (RP-HPLC). In the 10-mer RNA duplex model system a 4-isocyano TEMPO spin-label is individually attached to one strand and two strands are annealed to measure distances. This methodology is limited to systems in which two oligonucleotides are annealed together. To circumvent this limitation and also to explore single-strand dynamics a new methodology was implemented, double spin-labeling. Double spin-labeled single-stranded RNA was investigated as a single-strand and within a duplex via MALDI-TOF-MS, EPR spectroscopy and RP-HPLC. A double spin-labeling strategy in this work will be applicable to large complex RNAs like Group I intron of Tetrahymena thermophilia.

The function of biomolecules is very sensitive to structure. Folding in proteins and nucleic acids is a hierarchical process progressing from primary to secondary, then tertiary, and finally, quaternary structures. RNA in its folded form performs a variety of biological activities. Obtaining intramolecular distance measurements makes it possible to generate structural models along the folding pathway that may be related to the overall function of the molecule. Distances can be measured by Site-Directed Spin-Labeling (SDSL), in which nitroxyl spin-label probes are attached and observed by EPR spectroscopy. Spin-labels can provide information concerning structure and conformational changes because they are particularly sensitive to molecular motion and interspin distances. Continuous-wave EPR spectroscopy has been commonly applied to detect and monitor nitroxide spin-label probes within biological systems. A previous published SDSL study from this laboratory investigated a 10-mer RNA duplex model system with spin-label probe succinimdyl-2,2,5,5-tetramethyl-3-pyrroline-1-oxyl-carboxylate; however, an increased spin-labeling efficiency was observed with an isocyanate derivative of tetramethylpiperidyl-N-oxy (TEMPO). In this thesis, a 4-isocyano TEMPO spin-label probe replaced the previously used succinimdyl-2,2,5,5-tetramethyl-3-pyrroline-1-oxyl-carboxylate in 10-mer SDSL studies. The influence of labeling with the 4-iscocyano TEMPO spin-label in a 10-mer RNA model system was investigated with thermal denaturation, Matrix Assisted Laser Desorption Time of Flight Mass Spectrometry (MALDI-TOF-MS), Electron Paramagnetic Resonance (EPR) spectroscopy, and reverse phase high performance liquid chromatography (RP-HPLC). In the 10-mer RNA duplex model system a 4-isocyano TEMPO spin-label is individually attached to one strand and two strands are annealed to measure distances. This methodology is limited to systems in which two oligonucleotides are annealed together. To circumvent this limitation and also to explore single-strand dynamics a new methodology was implemented, double spin-labeling. Double spin-labeled single-stranded RNA was investigated as a single-strand and within a duplex via MALDI-TOF-MS, EPR spectroscopy and RP-HPLC. A double spin-labeling strategy in this work will be applicable to large complex RNAs like Group I intron of Tetrahymena thermophilia.

This thesis describes the synthesis of a range of novel dithiadiazolyl radicals. The structures of these compounds are discussed. The physical properties of several compounds have been investigated using EPR spectroscopy and magnetic susceptibility studies. Chapter one begins with an overview of the chemistry of 1,2,3,5-dithiadiazolyl radicals. A general discussion of the history of organic conductors and magnets, and the terms involved in some of the techniques used is given in order to provide a background to the work presented. The second chapter outlines the synthesis and general characterisation of all the dithiadiazolyl radicals discussed in this thesis. A proposed mechanism for the conversion of parent nitriles into dithiadiazolyl radicals has been included. The third chapter describes the solid state structures of three dichlorophenyl dithiadiazolyl derivatives (2,4-, 2,5- and 3,5-dichlorophenyl-l,2,3,5-dithiadiazolyl). A further polymorph of 3,5-dichlorophenyl-1,2,3,5-dithiadiazolyl has also been included. The magnetic susceptibility of 2,4- and 3,5-dichlorophenyl-1,2,3,5-dithiadiazolyl has been investigated and the EPR analysis of all three compounds has been performed. These compounds are the first examples of neutral dithiadiazolyl radicals that form evenly spaced, segregated stacks in the solid state. Chapter four describes the dimer stacking structures of two further dichlorophenyl dithiadiazolyl derivatives (2,3- and 3,4-dichlorophenyl-1,2,3,5-dithiadiazolyl). The fifth chapter discusses the association of 3,5-dibromphenyl-1,2,3,5-dithiadiazolyl in the solid state. An investigation of this compound by EPR spectroscopy is also presented. Chapter six describes the trans cofacial association of p-iodophenyl-1,2,3,5-dithiadiazolyl in the solid state, only the second published example of this mode of dimerisation. The synthesis of o- and p-iodobenzonitrile are also described. An investigation of the EPR signal of this compound has also been included. Chapter seven describes the specialised techniques used in the synthesis of all the compounds. A list of the instruments used for analysis is also included.

Natural occurring riboswitches, a class of RNA molecules, are able to control the implementation of genetic information through the regulation of gene expression. Both, natural and synthetic riboswitches, so-called aptamers, are complex folded structures, which bind specific ligands. Ligand binding in turn regulates gene expression. In the first section of this work continuous wave (cw) and pulse EPR spectroscopy in combination with site-directed spin labeling was performed to investigate the dynamics and conformational changes of the synthetic tetracycline (Tc) riboswitch. The results obtained herein indicate a thermodynamic equilibrium of two aptamer conformations in the absence of Tc, where one of these conformations is captured upon ligand binding. Aptamer structures have been modeled based on the two equilibrium conformations in the absence of the ligand, and on the captured aptamer conformation in the presence of Tc. All RNA models have been verified by MD simulations by comparing experimentally obtained interspin distances with simulated ones. The incorporation of DNA mutations such as nucleobase changes, double-strand breaks and mispairings can lead to structural and conformational changes of DNA domains which in turn are related to inheritable diseases, cancer and aging. Within this project, the copper(I)-catalyzed Huisgen-Sharpless-Meldal alkyne-azide cycloaddition (CuAAC) ‘click reaction’ was introduced as a powerful modification of existing spin labeling strategies. Interspin distances and exceptionally narrow distribution widths were determined by cw and pulse EPR experiments. The results of the MD simulations exhibit a very good agreement between simulated and experimentally obtained distances. Furthermore, the new spin labeling protocol was used to identify mismatch-induced conformational changes of spin labeled DNA. The application of cw and orientation selective pulse EPR measurements provided insights into the dynamics and structural alterations caused by mismatched base pairs. Interspin distances have been found to depend on the type and nearest neighbor environment of the mismatch. The changes are transferred to the base pairs carrying the spin labels.

Within the PhD. thesis the problems of H-transfer from selected types of phenols, aromatic secondary amines and compounds with acidic CH bond were investigated. This process was initiated by the action of redox agents, mostly PbO2, in nonpolar solvents. The abstraction of hydrogen atom results in the formation of radical intermediates, which were detected by EPR spectroscopy. Because in most cases, highly unstable radicals were produced, it was neccessary to apply the technique of indirect detection, so called spin trapping. This method enables to interpret the character of generated radicals on the basis of EPR parameters of radical adducts, which are formed by their reaction with suitably chosen compounds, spin traps, which are added to the reaction system. As spin traps, the aromatic nitrosocompounds were preferentially used. Due to the fact that EPR spectra of radical adducts were in most cases very complex, their interpretation was possible only using the computer simulation. Based on this approach, new information about the character of addition of phenoxyl tradicals to aromatic nitrosocompounds, stability of alkyl substituents in methyl substituted phenols and character of C-radicals, generated from substituted coumarines and beta-diketones, was obtained. Simultaneously, the small ability of nitrogen radicals, primary products of the splitting of NH bond, to enter the reaction with spin traps was proved by aromatic secondary amines.

Intracellular protein-protein interactions (PPIs) play a vital role in many biological processes. Although they are viewed as of high biological interest they prove difficult to explore as potential targets for drug discovery. Numerous studies have shown α- helical peptides 'locked' in their respective bioactive structure can greatly increase their performance by increasing their target affinity, resistance to proteolysis as well as facilitating cellular uptake. A striking feature of literature to date is how few studies utilise different stapling techniques when developing inhibitors for PPIs. Current methods generally exploit ruthenium catalysed ring closing metathesis (RCM) or copper catalysed alkyne/azide click (CuAAC) chemistry to generate geometrically constrained peptides. Even though these methods have shown great potential they both share a fundamental limitation as the chemistry can only be employed on small synthetic peptides and cannot be extended to larger proteins. Thiol-ene coupling (TEC) chemistry (Chapter 1) which is often described as a 'click' reaction due to its fast reaction rates, high yields, wide functional group tolerance and insensitivity to ambient oxygen and water has the potential to solve this challenge. Thiol-ene chemistry was investigated as an alternative stapling strategy by employing the naturally occurring amino acid L-cysteine (Cys) as a source of the thiyl radical and L-homoallylglycine (Hag), a non-natural amino acid shown to act as a methionine surrogate in protein synthesis to act as a source of an alkene functionality to form a potentially expressible thioether tether in Chapter 2. However, due to unsatisfactory results from the intramolecular thiol-ene cyclisation at the molar concentrations required for peptide or protein modification, and a promising new lead, the closely related thiol-yne reaction was investigated as an alternative in Chapter 3. Using a small library of peptides (14 mers) derived from α-Synuclein (αSyn), a protein mainly found in the presynaptic terminals in the brain and is believed to be key to the pathological progress of Parkinson's disease, a successful macrocyclisation was achieved between the side chains of cysteine (Cys) and homopropargylglycine (Hpg). Although the vinyl-thioether tether did not confer any helical conformation on the stapled peptides, the results clearly demonstrate a potential route for the development of expressible staples. Electron paramagnetic resonance (EPR) spectroscopy in combination with site-directed spin labelling (SDSL) of biomolecules has become a powerful tool for studying the structure and conformational dynamics of biomolecules. Typically, proteins are modified in a site-specific manner by utilising the side chains of cysteine residues to form disulphide bonds with spin active compounds, however, this strategy has its limitations. In Chapter 3 thiol-ene chemistry was investigated as an alternative biorthogonal method to spin label proteins and peptides. The newly synthesised sulfhydryl bearing nitroxide spin label was found to degrade upon exposure to radical promoting conditions, however, an alternative strategy was explored using more classical thiol-Michael chemistry to spin label dehydroalanine (Dha) modified peptides giving the desired spin labelled complex.

No description available.

Currently, there is interest in the development of molecular-scale devices for use in quantum information processing (QIP). With this application in mind, physical studies on antiferromagnetically coupled molecular wheels [Cr7MF3(Etglu)(O2CtBu)15(phpy)], where M is a divalent metal cation (M = Mn2+, Zn2+, Ni2+) have been pursued. The heterometallic wheels contain an octagon of metal centres, which are bridged by fluoride ions, pivalate groups and a chiral N-ethyl-D-glutamine molecule which is penta-deprotonated and bound to the metal sites through all available O-donors. They are deep purple in colour and they have been named purple-Cr7M. There is antiferromagnetic coupling between adjacent metal centres, J » -8 cm-1, resulting in a non-zero net spin ground state. The spin-Hamiltonian parameters of this family have been determined.At the heterometal site of purple-Cr7M wheels there is a terminal ligand which can be substituted for a variety of N-donor organic ligands. A series of bidentate N-donor linkers has been used to link Cr7Ni wheels (each wheel Seff = 1/2) to create prototype two-qubit systems. Multi-frequency EPR spectroscopy and SQUID magnetometry has been used to extract the spin-Hamiltonian parameters of this family. It has been shown that the single wheels can be linked together electronically as well as chemically. It has been found that for the unsaturated linkers, there is a weaker interaction between Cr7Ni wheels when longer linkers are used. The strength of interaction is smaller for the saturated linkers than for the unsaturated linkers.The formation of 'green'-Cr7M wheels is different, being templated around a cation. Two new types of wheels have been studied: [tBuCONHC6H12NH2C6H12NHCOtBu][Cr7M2+F8(O2CtBu)16] and [Cs?Cr7MF8(O2CtBu)16]·0.5MeCN (where, M = Mn2+, Zn2+, Ni2+), where the former is templated around a long dialkylammonium group and the latter around a caesium cation. The effect of the templating cation on spectroscopic properties has been determined.Physical studies on a family of antiferromagnetically-coupled homometallic clusters have been pursued. They consist of cyclic arrays of homometallic Cr3+ ions in either a octametallic wheel or hexametallic horseshoes. The horseshoes have the general formula: [CrxFx+5L2x-2]n3- (where L = carboxylate). Cr3+ centres are bridged by pivalate groups and fluorides, while Cr3+ centres at the ends of the chain have terminal fluorides completing their coordination sphere. These terminal fluoride groups are labile enough to be substituted, e.g. [EtNH2][Cr6F7(O2CtBu)10(acac)2] is the product of a substitution reaction with acetylacetone.

In the framework of Ph.D. thesis the evaluation of radical reactions of four groups of secondary amines R1–NH–R2 in the presence of some selected types of agents was performed using EPR spectroscopy. First group was represented by peroxoagents (3-chloroperbenzoic acid, tBuO2 radicals), the second group involved compounds of PbIV+ (PbO2, Pb(OAc)4) functioning as hydrogen-abstracting agents. In the presence of peroxyagents the formation of corresponding aminoxyl radicals R1–NO–R2 was demonstrated. In the case of surplus of tBuO2 radicals these radicals enter the consecutive reactions which products are new types of secondary aminoxyls. By the interpretation of reaction mechanism the intermediary formation of nitrones was assumed. These function in later steps of the reaction as spin traps for tBuO2 radicals. By the study of radical mechanism of amines R1–NH–R2 initiated by PbIV+ agents was proved, that these compounds hydrogen atom not only from –NH– group under formation of aminyl radicals, but also from C–H bonds in substituents R1, R2 (the formation of C-radicals). Because both groups of the radicals formed are characterized by high reactivity, their identification was possible only using spin-trapping method. In the course of the investigation of radical reactions of secondary amines the big amount of experimental EPR spectra was obtained. These exhibited in many cases very high hyperfine structure. Due to this fact it was necessary to perform the detailed evaluation on the basis of spectral simulation. Except of some special cases using this approach the interpretation of majority of registered EPR spectra was interpreted and EPR parameters were summarized in tables.