Bacterial auto-nemesis: templating polymers for cell sequestration

The detection and control of microorganisms such as bacteria is important in a wide range of industries and clinical settings. Detection, binding and removal of such pathogenic contaminants can be achieved through judicious consideration of the targets which are available at or in the bacterial cell. Polymers have the ability to present a number of binding ligands for cell targeting on one macromolecule and so avidity of interaction can be greatly increased. The goal of the project was to test whether polymers generated with bacteria in situ would have their composition significantly altered to determine if a templating process was occurring. It was also anticipated that the templated polymers would have better re-binding properties than those produced in the absence of bacteria. A series of chemical functionalities were analysed for their binding properties to bacteria. The functionalities were chosen with consideration to the cell surface characteristics. Further to identification of the most binding and least binding functionalities the polymers were tested for their cytotoxicity against bacteria and human epithelial cells. Concentration ranges were determined which could facilitate bacterial binding and templating yet minimise the lethality of the processes. Templated polymers of the bacteria were generated using a novel method of atom transfer radical polymerisation (ATRP) which we have termed bacterial activated atom transfer radical polymerisation (b-ATRP). This polymerisation method has maximised the potential for templating processes to occur during the polymerisation. Templated polymers differed in both their composition and their binding behaviour to non-templated polymers. The bacterial organic reduction process has also been demonstrated to have greater scope for use within the organic chemistry field as demonstrated by the use of this system to enable in "click-chemistry" via the reduction of copper

Making silicone rubber highly resistant to bacterial attachment using thiol-ene grafting

Biomedical devices are indispensable in modern medicine yet offer surfaces that promote bacterial attachment and biofilm formation, resulting in acute and chronic healthcare-associated infections. We have developed a simple method to graft acrylates to silicone rubber, polydimethylsiloxane (PDMS), a commonly used device material that is often colonized by bacteria. We demonstrate a novel method whereby nontoxic bacteria attachment-resistant polymers can be readily grafted from and grafted to the surface using thiol-ene chemistry, substantially reducing bacterial colonization. With use of this approach, bacterial biofilm coverage can be reduced by 99% compared with standard PDMS in an in vitro assay. This grafting approach offers significant advantages over commonly used physisorbed coatings, especially in areas of high shear or mechanical stress. Furthermore, the approach is versatile such that the grafted material properties can be tailored for the desired final application

Conformational Heterogeneity in a Fully Complementary DNA Three-Way Junction with a GC-Rich Branchpoint.

DNA three-way junctions (3WJs) are branched structures that serve as important biological intermediates and as components in DNA nanostructures. We recently derived the global structure of a fully complementary 3WJ and found that it contained unpaired bases at the branchpoint, which is consistent with previous observations of branch flexibility and branchpoint reactivity. By combining high-resolution single-molecule Förster resonance energy transfer, molecular modeling, time-resolved ensemble fluorescence spectroscopy, and the first (19)F nuclear magnetic resonance observations of fully complementary 3WJs, we now show that the 3WJ structure can adopt multiple distinct conformations depending upon the sequence at the branchpoint. A 3WJ with a GC-rich branchpoint adopts an open conformation with unpaired bases at the branch and at least one additional conformation with an increased number of base interactions at the branchpoint. This structural diversity has implications for branch interactions and processing in vivo and for technological applications

Design of photoactivatable metallodrugs : selective and rapid light-induced ligand dissociation from half-sandwich [Ru([9]aneS3)(N–N′)(py)]2+ complexes

The synthesis of the inert Ru(II) half-sandwich coordination compounds, [Ru([9]aneS3)(bpy)(py)][PF6]2 (1, [9]aneS3 = 1,4,7-trithiacyclononane, bpy = 2,2′-bipyridine, py = pyridine), [Ru([9]aneS3)(en)(py)][PF6]2 (2, en = 1,2-diaminoethane), and [Ru([9]aneN3)(en)(dmso-S)][PF6]2 (3, [9]aneN3 = 1,4,7-triazacyclononane), is reported along with the X-ray crystal structure of 1. We investigated whether these complexes have photochemical properties which might make them suitable for use as pro-drugs in photochemotherapy. Complexes 1 and 2 underwent rapid (minutes) aquation with dissociation of the pyridine ligand in aqueous solution when irradiated with blue light (λ = 420 or 467 nm). The photodecomposition of 3 was much slower. All complexes readily formed adducts with 9-ethylguanine (9-EtG) when this model nucleobase was present in the photolysis solution. Similarly, complex 1 formed adducts with the tripeptide glutathione (GSH), but only when photoactivated. HPLC and MS studies of 1 showed that irradiation promoted rapid formation of 1:1 (major) and 1:2 (minor) adducts of the oligonucleotide d(ATACATGCTACATA) with the fragment {Ru([9]aneS3)(bpy)}2+. Density functional theory (DFT) calculations and time-dependent DFT reproduced the major features of the absorption spectra and suggested that the lowest-lying triplet state with 3MLCT character, which is readily accessible via intersystem crossing, might be responsible for the observed dissociative behavior of the excited states. These complexes are promising for further study as potential photochemotherapeutic agents

Metallation‐induced heterogeneous dynamics of DNA revealed by single‐molecule FRET

The metallation of nucleic acids is key to wide‐ranging applications, from anticancer medicine to nanomaterials, yet there is a lack of understanding of the molecular‐level effects of metallation. Here, we apply single‐molecule fluorescence methods to study the reaction of an organo‐osmium anticancer complex and DNA. Individual metallated DNA hairpins are characterized using Förster resonance energy transfer (FRET). Although ensemble measurements suggest a simple two‐state system, single‐molecule experiments reveal an underlying heterogeneity in the oligonucleotide dynamics, attributable to different degrees of metallation of the GC‐rich hairpin stem. Metallated hairpins display fast two‐state transitions with a two‐fold increase in the opening rate to ~2 s‐1, relative to the unmodified hairpin, and relatively static conformations with long‐lived open (and closed) states of 5 s to ≥ 50 s. These studies show that a single‐molecule approach can provides new insight into metallation‐induced changes in DNA structure and dynamics

Photoactivated chemotherapy (PACT) : the potential of excited-state d-block metals in medicine

The fields of phototherapy and of inorganic chemotherapy both have long histories. Inorganic photoactivated chemotherapy (PACT) offers both temporal and spatial control over drug activation and has remarkable potential for the treatment of cancer. Following photoexcitation, a number of different decay pathways (both photophysical and photochemical) are available to a metal complex. These pathways can result in radiative energy release, loss of ligands or transfer of energy to another species, such as triplet oxygen. We discuss the features which need to be considered when developing a metal-based anticancer drug, and the common mechanisms by which the current complexes are believed to operate. We then provide a comprehensive overview of PACT developments for complexes of the different d-block metals for the treatment of cancer, detailing the more established areas concerning Ti, V, Cr, Mn, Re, Fe, Ru, Os, Co, Rh, Pt, and Cu and also highlighting areas where there is potential for greater exploration. Nanoparticles (Ag, Au) and quantum dots (Cd) are also discussed for their photothermal destructive potential. We also discuss the potential held in particular by mixed-metal systems and Ru complexes

Bacteria-instructed synthesis of polymers for self-selective microbial binding and labelling

The detection and inactivation of pathogenic strains of bacteria continues to be an important therapeutic goal. Hence, there is a need for materials that can bind selectively to specific microorganisms, for diagnostic or anti-infective applications, but which can be formed from simple and inexpensive building blocks. Here, we exploit bacterial redox systems to induce a copper-mediated radical polymerisation of synthetic monomers at cell surfaces, generating polymers in situ that bind strongly to the microorganisms which produced them. This ‘bacteria-instructed synthesis’ can be carried out with a variety of microbial strains, and we show that the polymers produced are self-selective binding agents for the ‘instructing’ cell types. We further expand on the bacterial redox chemistries to ‘click’ fluorescent reporters onto polymers directly at the surfaces of a range of clinical isolate strains, allowing rapid, facile and simultaneous binding and visualisation of pathogens

The impact of surface chemistry modification on macrophage polarisation

Macrophages are innate immune cells that have a central role in combating infection and maintaining tissue homeostasis. They exhibit remarkable plasticity in response to environmental cues. At either end of a broad activation spectrum are pro-inflammatory (M1) and anti-inflammatory (M2) macrophages with distinct functional and phenotypical characteristics. Macrophages also play a crucial role in orchestrating immune responses to biomaterials used in the fabrication of implantable devices and drug delivery systems. To assess the impact of different surface chemistries on macrophage polarisation, human monocytes were cultured for 6 days on untreated hydrophobic polystyrene (PS) and hydrophilic O2 plasma-etched polystyrene (O2-PS40) surface. Our data clearly show that monocytes cultured on the hydrophilic O2-PS40 surface are polarised towards an M1-like phenotype, as evidenced by significantly higher expression of the pro-inflammatory transcription factors STAT1 and IRF5. By comparison, monocytes cultured on the hydrophobic PS surface exhibited an M2-like phenotype with high expression of mannose receptor (MR) and production of the anti-inflammatory cytokines IL-10 and CCL18. While the molecular basis of such different patterns of cell differentiation is yet to be fully elucidated, we hypothesise that it is due to the adsorption of different biomolecules on these surface chemistries. Indeed our surface characterisation data show quantitative and qualitative differences between the protein layers on that the O2-PS40 surface compared to PS surface which could be responsible for the observed differential macrophage polarisation on each surface

Considerations on Genre and Gender Conventions in Translating from Old English

The Old English poem The Wife's Lament is an extremely conventional and, at the same time, original text. It portrays a female character suffering for the absence of her loved one, through the framework of the so-called 'elegiac' style and a mainly heroic vocabulary. The traditional exile theme is, thus, interwoven with the uncommon motif of love sickness. While this appraisal of the poem is the most widely accepted one, disagreement still remains about the translation of some keywords, strictly related to the exile theme, such as sīþ or wræcsīþ. The aim of this paper is to examine diverging readings and glosses of the above mentioned 'exilic/elegiac' keywords, and to show that an accurate translation should not neglect a thorough appraisal of the text in its complexity and the association with related literary patterns and imagery in other poetic and prose texts

Bacterial auto-nemesis - templating polymers for cell sequestration

The detection and control of microorganisms such as bacteria is important in a wide range of industries and clinical settings. Detection, binding and removal of such pathogenic contaminants can be achieved through judicious consideration of the targets which are available at or in the bacterial cell. Polymers have the ability to present a number of binding ligands for cell targeting on one macromolecule and so avidity of interaction can be greatly increased. The goal of the project was to test whether polymers generated with bacteria in situ would have their composition significantly altered to determine if a templating process was occurring. It was also anticipated that the templated polymers would have better re-binding properties than those produced in the absence of bacteria. A series of chemical functionalities were analysed for their binding properties to bacteria. The functionalities were chosen with consideration to the cell surface characteristics. Further to identification of the most binding and least binding functionalities the polymers were tested for their cytotoxicity aga.inst bacteria and human epithelial cells. Concentration ranges were determined which could facilitate bacterial binding and templating yet minimise the lethality of the processes. Templated polymers of the bacteria were generated using a novel method of atom transfer radical 27 polymerisation (ATRP) which we have termed bacterial activated atom transfer radical polymerisation (b-ATRP). This polymerisation method has maximised the potential for templating processes to occur during the polymerisation. Templated polymers differed in both their composition and their binding behaviour to non-templated polymers. The bacterial organic reduction process has also been demonstrated to have greater scope for use within the organic chemistry field as demonstrated by the use of this system to enable in "click-chemistry" via the reduction of copper. 28EThOS - Electronic Theses Online ServiceGBUnited Kingdo