Routes to mononuclear Os(II) and Os(0) cluster complexes containing P-donor ligands (PR3, PP) were studied using the osmium salts Y2[OsX6] (Y = NH4, K; X = Cl, Br, I) as precursors. Carbonylation of the precursors and then subsequent reaction with tertiary phosphines afford neutral complexes of the type cis,cis,trans-[OsX2(CO)2(PR3)2] (X = Cl, Br, I, PR3 = tertiary phosphine), which were characterized by combination of infrared and Raman spectroscopy, NMR (1H, 13C, 31P) spectroscopy and elemental analysis. The compounds cis,cis,trans-[OsBr2(CO)2{Ph2P(CH2C6H5)}2] (2b), cis,cis,trans-[OsBr2(CO)2{P(CH2C6H5)}3] (3b) and cis,cis,trans-[OsBr2(CO)2{P(C6H11)}2] (4b) have been further characterized by single crystal X-ray diffraction. Thermogravimetric analysis of these complexes shows multiple stages of decomposition that can be directly related to the loss of ligands.Microwave-promoted reaction of the osmium salt precursor with bidentate phosphine ligands affords the neutral complexes [OsX2(PP)2] (X = Cl, Br, I; PP = bidentate phosphine ligand). A stepwise substitution pathway has been suggested resulting in the formation of compounds cis/trans-[OsX2(PP)2] in which the ligands display different reactivity. The size of the bidentate phosphine ligand determines the stereochemistry of the product complex. Reaction with short ligands such as dppm selectively yield the cis-[OsX2(PP)2] isomers. Whilst reaction with dppe and dppp under the same conditions gives the trans-[OsX2(PP)2] isomers. Steric bulkiness of the ligands influences the stereochemistry. The cis-[OsX2(PP)2] isomers are thermally stable with onset of decomposition at temperatures above 300 °C. In contrast, the trans-[OsX2(PP)2] isomers were found to be thermally less stable and found to decompose in the range 195 – 251 °C. Microwave irradiation of [Os3(CO)12] in the presence of PPh3 in acetonitrile solution gave a mixture of disubstituted and trisubstituted phosphine cluster complexes which have been characterized by IR, Raman, 1H and 31P (where applicable) NMR spectroscopies. Variation of solvents influences substitution and give a mixture of monosubstituted and disubstituted phosphine cluster. Complexes of the type [OsX2(PP)2] exhibited both anticancer and antimicrobial activity. In some instances the activity was comparable to that of the reference drugs. The complexes cis,cis,trans-[OsX2(CO)2(PR3)2] and phosphine substituted derivatives of [Os3(CO)12] exhibited moderate antibacterial activity against Gram-negative bacterial strains and some fungal strains under study. / Chemistry / M. Sc. (Chemistry)

Cis-diamminedichloroplatinum (II), or cisplatin, is a widely prescribed anticancer compound, currently one of only three platinum (II) complexes FDA approved for cancer treatment. Despite its widespread use, we lack a comprehensive picture of global drug targets, which would lend valuable insights into the molecular mechanisms of action and resistance in different tissues. Drug binding to genomic DNA is an accepted cause of downstream apoptotic signaling, but less than 10% of Pt (in the case of cisplatin) accumulates within genomic DNA. Non-genomic contributions to cisplatin's therapeutic action are also under active investigation. In particular, cisplatin treatment can disrupt RNA-based processes such as splicing and translation. Pt(II) targeting of non-DNA species such as RNA may contribute to or sensitize a cell to the downstream effects of this drug, including the induction of apoptosis. Chapter I summarizes the activity profile of Pt(II) therapeutics, describing cellular uptake, cellular localization, incidences of Pt(II) accumulation within RNA, and RNA processes affected following drug treatment. Chapter II reports our thorough investigation of the distribution of Pt species throughout messenger and ribosomal RNA, with the discovery that Saccharomyces cerevisiae ribosomes act as a de facto cellular Pt sponge. In Chapter III, we report the synthesis of an azide-functionalized platinum (II) species, picazoplatin, for post-treatment click labeling and isolation of drug targets in vivo. Picazoplatin was designed to circumvent mislocalization and misprocessing of Pt typically encountered when trying to track small molecules tethered to large, charged fluorophores. This chapter contains several proof-of-principle studies validating the use of this class of reagents for future purification and sequencing of Pt-bound nucleic acids. Chapter IV describes the first application of the click-capable Pt reagent technology: the demonstration of significant in-gel fluorescent detection of Pt-bound ribosomal RNA and transfer RNA extracted from picazoplatin-treated S. cerevisiae and the first evidence that cellular tRNA is a platinum substrate. Chapter V summarizes these data, which suggest a potential ribotoxic mechanism for cisplatin cytotoxicity and broadly describe a convenient click chemistry methodology that can be applied to identify other metal or covalent modification-based drug targets. This dissertation includes previously published and unpublished co-authored material.

Yes / This report presents a new library of organometallic iridium(III) compounds of the type [Cp*IrCl(L)] (Cp*=pentamethylcyclopentadienyl and L=a functionalized β‐ketoiminato ligand) showing moderate to high cytotoxicity against a range of cancer cell lines. All compounds show increased activity towards colorectal cancer, with preferential activity observed against the immortalized p53‐null colorectal cell line, HCT116 p53‐/‐, with sensitivity factors (SF) up to 26.7. Additionally, the compounds have excellent selectivity for cancerous cells when tested against normal cell types, with selectivity ratios (SR) up to 35.6, contrary to that of cisplatin, which is neither selective nor specific for cancerous cells (SF=0.43 and SR=0.7–2.3). This work provides a preliminary understanding of the cytotoxicity of iridium compounds in the absence of p53 and has potential applications in treatment of cancers for which the p53 gene is absent or mutant.

No / Herein we report a library of new ruthenium(II) complexes which incorporate a range of functionalised β -ketoiminate ligands. The complexes undergo an unusual reduction from Ru(III) to Ru(II), and consequently incorporate carbonyl ligands from the 2-ethoxyethanol solvent, forming ruthenium dicarbonyl complexes. In order to address the potential applications of these complexes, we have screened the library against a range of tumour cell lines, however, all compounds exhibit low cellular activity and this is tentatively assigned to the decomposition of the compounds in aqueous media. Studies to establish the antifungal and antibacterial potential of these complexes was addressed and show increased growth inhibitions for C. neoformans and S. aureus species. / The authors would like to thank the universities of Leeds, Huddersfield and Bradford for internal financial support.

IsdG and IsdI are non-canonical heme oxygenases (HO) from Staphylococcus aureus that catalyze the oxidative cleavage of heme to give novel organic products (staphylobilins) and iron as a nutrient for the pathogen. Comparison of the reported equilibrium dissociation constant (Kd) values for heme from IsdG and IsdI compared to the reported concentration of the labile heme pool called into question whether these enzymes are competent HOs in vivo. We took advantage of a second-sphere Trp whose fluorescence is quenched upon heme binding, which led to Kd values 2-3 orders of magnitude smaller than reported in the literature. Importantly, these Kd values were on the same order of magnitude as human HO, precluding design of a competitive inhibitor as an effective therapeutic. Based upon the kinetic and equilibrium data, and the finding that the half-life of IsdG is increased 2.5-fold by the presence of heme, we proposed IsdG is the main HO involved in iron acquisition which motivated further characterization of IsdG. IsdG-catalyzed heme catabolism proceeds through ferric-peroxoheme and meso-hydroxyheme intermediates en route to staphylobilin. A second-sphere Asn is known to be critical for enzymatic function, but its role in heme cleavage was unknown. Site-directed mutagenesis was employed to probe the role of Asn using ferric-azidoheme and ferric-cyanoheme as models of the putative ferric-peroxoheme intermediate. An optical spectroscopic study established that a hydrogen-bond between Asn and the iron-ligating (α) atom of the distal ligand perturbs the heme electronic structure. Density functional theory (DFT) suggested this hydrogen-bond triggers rotation of the distal ligand, which was corroborated by circular dichroism (CD), and delocalizes spin density onto the meso carbons. Electron paramagnetic resonance (EPR) revealed the Asn hydrogen-bond increases the Fe 3dxy character in the singly occupied molecular orbital (SOMO), a mechanism that can increase spin density on the meso carbons. Finally, the Asn hydrogen-bond moves the meso carbon resonances downfield in the 13C nuclear magnetic resonance (NMR) spectrum, consistent with excess spin density, confirming a DFT-predicted, Asn-induced spin delocalization. These results suggest IsdG funnels the reactivity of ferric-peroxoheme toward heme hydroxylation through an Asn-dependent bridged transition state, circumventing production of reactive, uncontrolled intermediates.

acase@tulane.edu

<p>In this dissertation several metalloproteins, both metal transport proteins and the classic metalloprotein hemoglobin, are investigated using a variety of biophysical and electrochemical techniques. In each case, thermodynamic measurements provide insight into the role and mode of action of the metalloprotein under investigation. In Chapters 2 and 3, we focus on the thermodynamic properties of the metal while bound by the protein. In Chapter 4, we focus on the thermodynamic properties of the protein with and without the metal. In Chapter 5, we utilize both the metal and the protein as our probe.</p> <p>In Chapter 2, we probe the thermodynamic properties of the heme-bound iron to elucidate the structure-function relationships underlying two important physiological responses of hemoglobin (Hb): the Root Effect of hemoglobin from certain fish and the different nitrite reactivities of hemoglobins from clams. Hemoglobins of some fish exhibit significantly lowered oxygen affinity at low pH, allowing for proton-mediated release of O₂. This phenomenon, known as the Root Effect, serves as a proton-driven pump delivering O₂ to the swim bladders and eyes of the fish. The clam, ,<italic>L. pectinata</italic>, expresses functionally distinct Hb I that transports H₂S and Hb II that transports O₂. These two hemoglobins differ widely in their reactivity with nitrite, a reactant of great importance to the study of vasodilation in humans. The structural basis of the extreme pH-sensitivity of the Root Effect Hbs and the extreme reactivities of the <italic>Lucina Hbs</italic> with nitrite are debated. Focusing on the metal as the probe, we investigate the reduction potentials of these Hbs using spectroelectrochemistry and compare our findings with oxygen binding studies performed by our collaborators. In both cases, our data strongly suggest that steric hindrance is the determining factor governing the respective physiological response of each hemoglobin. </p> <p>In Chapter 3, we again use the metal as the probe to determine the reduction potential of titanium bound by transferrin (Tf). Tf is the human iron transport protein that can also bind titanium. To address the possible mechanisms of titanium transport through the hypothesized redox-mediated Fe₂-Tf transport pathway, a modified spectroelectrochemistry (SEC) method was developed to measure the electrochemical properties of metalloproteins with very negative potentials. However, the reduction potential of Ti₂-Tf is far too negative to access with our system. As an alternative approach, the redox properties of several model titanium and iron compounds were characterized in order to develop a linear free energy relationship (LFER) allowing us to estimate the reduction potential of Ti₂-Tf to be ca. -900 mV vs. NHE. Our results indicate that the reduction potential of Ti₂-Tf is too low to be reduced by biological reducing agents and suggest that transferrin-mediated titanium transport follows a different mechanism than iron transport.</p> <p>In Chapter 4, our focus shifts to the thermodynamic properties of the protein. Some pathogenic Gram-negative bacteria such as <italic>N. gonorrhoeae</italic> steal iron from their human host by expressing a receptor (TbpA/TbpB), which binds the human iron transport protein transferrin (Tf). Once iron crosses the outer membrane, ferric binding protein (FbpA) transports it across the periplasm to the cytosol. Focusing on the protein, we investigated the protein-protein interactions involved in this transport process and the roles that TbpA and TbpB play with the use of an H/D exchange and mass spectrometry based method termed SUPREX. We report herein the first direct measurement of periplasmic FbpA binding to the outer membrane protein TbpA and we demonstrate that both TbpA and TbpB individually can deferrate Tf without energy supplied from TonB, resulting in sequestration by apo-FbpA.</p> <p>In Chapter 5, we extend our investigation of the <italic>N. gonorrhoeae</italic> iron uptake system by using the metal as the probe in one case and the protein as the probe in another case. TbpA, the &#946;-barrel receptor protein that is required for utilization of Fe₂-Tf as an iron source, has a plug domain which we hypothesize binds iron and interacts with FbpA on the periplasmic side of the outer membrane. Utilizing SUPREX to monitor the thermodynamic properties of protein folding, we investigate 1) the possible interactions between the TbpA-plug and FbpA and 2) the ability of the TbpA-plug to bind iron. </p> <p>Focusing on the metal as the probe, we designed an experimental apparatus to investigate the possible thermodynamic effects of the TbpA/TbpB receptor on the release of iron from Tf. We report the use of a competitive iron chelator and equilibrium dialysis allows for the spectroscopic monitoring of iron release from Tf in the absence of FbpA, but in the presence of opaque bacterial membrane preparations containing the receptor.</p> / Dissertation

In this thesis, studies of the nature of different transition metal-hydride complexes are described. The first part deals with the enantioswitchable behaviour of rhodium complexes derived from amino acids, applied in asymmetric transfer hydrogenation of ketones. We found that the use of amino acid thio amide ligands resulted in the formation of the R-configured product, whereas the use of the corresponding hydroxamic acid- or hydrazide ligands selectively gave the S-alcohol. Structure/activity investigations revealed that the stereochemical outcome of the catalytic reaction depends on the ligand mode of coordination. In the second part, an Fe hydrogenase active site model complex with a labile amine ligand has been synthesized and studied. The aim of this study was to find a complex that efficiently catalyzes the reduction of protons to molecular hydrogen under mild conditions. We found that the amine ligand functions as a mimic of the loosely bound ligand which is part of the active site in the hydrogenase. Further, an Fe hydrogenase active site model complex has been coupled to a photosensitizer with the aim of achieving light induced hydrogen production. The redox properties of the produced complex are such that no electron transfer from the photosensitizer part to the Fe moiety occurs. In the last part of this thesis, the development of a protocol for the transfer hydrogenation of ketones to secondary alcohols without the involvement of transition metal catalysts is described. A variety of ketones were efficiently reduced in 2-propanol using catalytic amounts of alkali alkoxide under microwave irradiation.

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