VictorJuiz/thesis huggingface dataset:
Ion channel crystallisation for structural studies

Ion channels control the movement of ions allowing specific ions to pass across biological membranes. This apparently simple process is used to control many essential physiological processes including the generation and propagation of neuronal excitation, regulation of insulin secretion, heartbeat and cell volume. Ion channels are able to do this by having multiple means of controlling the opening and closing of the ion conduction pathway. Several recent ion channel structures have shed light on the important process of ion selectivity as well as providing clues on channel gating. However, it is not yet completely clear how ion channels control the passage of ions across the membrane. My work is centred on understanding the gating process through structural analysis mostly using X-ray crystallography. In order to get to this point, crystals of the target ion channels need to be grown. This thesis presents four projects that were aimed at developing the techniques for over-expression, purification and crystallisation of ion channels or transporters making them amenable for structural studies.
One of the difficult steps in the structure determination pathway is obtaining well-ordered crystals of the target membrane protein. Crystals of the EcCLC channel (now found to be a transporter) from E. coli were initially unacceptable for structure determination. It was shown that the process of dehydration improved the quality and diffraction limit of the EcCLC crystals. This simple method provides a possible procedure for improving the resolution limit of soluble and membrane protein crystals.
Inward rectifier potassium channels (Kir channels) are a distinct subfamily that preferentially allows K+ ions to move into the cell. Two bacterial homologues of Kir channels, KirBac1.1 and KirBac3.1, were crystallised in 3-D and 2-D forms, respectively. As the crystal structure revealed the full-length model provided an excellent starting point for understanding the gating process in these channels.
The 3-D structure of KirBac1.1 was captured in the closed state. This K+ channel prevents ion movement by using hydrophobic residues to occlude the ion conduction pathway, altering the conformation of selectivity filter, misaligning the pore helices and decreasing the volume of the cavity.
The 2-D projection maps of KirBac3.1 were obtained in both closed and a potentially open conformations. These two maps provided evidence as to how the structural elements of the inner, outer, slide helices and the C-terminal domains change relative to one another as the channel moves between these two gating states. These motions were modelled as rigid body movements. They included bending of the inner helices, a rotation of the outer helices as they straighten in the membrane, translocation of the slide helices and C-terminal domain rotations.
It is hoped that the information gained from these projects, the plasmids construction, the expression systems and the protocols that were established for crystallisation of the ion channels and transporter will provide guidelines for future structural studies of membrane proteins.
Cenozoic deformation in a plate-boundary zone, Marlborough, New Zealand

The Marlborough Fault System is a zone of dextral transpression in continental crust at the southern end of the Hikurangi subduction system between the obliquely convergent Australian and Pacific plates. Detailed mapping of an area of deformed Tertiary cover sequence on the down thrown side of the Kekerengu Fault (the Kekerengu- Washdyke study area) has revealed two phases of deformation, De and Dl.
In the study area De consists of nine kilometre scale thrust faults cutting sediments derived from extra- and intra-basinal uplift. The timing of this episode is constrained by the age of the first clastic deposits and by a previously unidentified unconformity in this area of Late Miocene age. A clear sequence of Dl events is recognized deforming all earlier structures including Pliocene aged sediments. Although elsewhere in Marlborough a regional post-Pliocene ca. 20° clockwise block rotation has been previously identified, in the Kekerengu-Washdyke study area one site suggested no post-Pliocene rotation and another a large ca. 100° clockwise rotation. This lack of Dl rotation was unexplained and the large rotation attributed to localized Early Miocene deformation. Palaeomagnetic work carried out in this study has identified six more sites which contain this large rotation (average ca. 118+11°). The rotation therefore appears to be a regional event, likely to be a result of the location of Marlborough in the hinge zone at the southern end of the Hikurangi Margin. One site from dykes in basement rocks does not record this large rotation, indicating that the rotation occurred in upper layers detached from unrotating rock below by an unknown structure (such as a thrust fault), or that the rotation did not occur in this area. The large rotation is believed to have been achieved by pinning of the De thrusts to the south of the Marlborough region. The data suggests that the De thrusts in Marlborough were initially NW-trending and seaward, not landward-directed as was previously supposed. Palaeomagnetic work has also added to the evidence for a lack of Dl regional rotation on the downthrown side of the Kekerengu Fault. A previously unidentified second phase of Dl folding and 'bending' within the study area appears to have accommodated the regional rotation and suggests that the Kekerengu Fault acted as the eastern boundary of the Dl rotating block.
Awareness, action and feedback in domestic energy use

The nature of gas and electricity and the methods of distribution, billing and payment all contribute to the 'invisibility' of much domestic energy consumption in industrial and post-industrial societies. For the householder, understanding how to invest and behave in ways that will give affordable comfort with minimum environmental impact involves making sense of a hidden set of processes. This poses a major challenge, one that a range of energy advice programmes is attempting to meet. The main focus to date has been on the actions taken as a result of advice, with little attention paid to teaching and learning processes or to context. This thesis explores formal and informal processes by which householders learn about their energy use in order to develop a theoretical framework.
Constructivist learning theory guides the investigation and a variant of the 'conscious competence' model of learning is used as a starting point. The concept of'tacit knowledge' (foundational knowledge, usually acquired informally) is used in tracing the development of energy literacy. Empirical data come from householder surveys and from interviews of householders and advisers in five contrasting locations in the UK. Interpretation of this material demonstrates the construction of meaning through experience and interaction with others, and the potential role of the energy adviser as a trusted and knowledgeable person.
The building of tacit knowledge is crucial to the development of energy literacy and the householder's ability to absorb and evaluate new information. Energy advisers need to be able to identify and develop existing knowledge, and to form effective networks with social welfare programmes. The need to build awareness by following up advice wherever possible is stressed. There also needs to be a supportive learning infrastructure that includes easily accessible feedback on consumption, and the availability of accurate information for those who are knowledgeable and confident enough to teach themselves.
Adolescent dating violence and the negotiation of gender

The purpose of the thesis is to investigate psychological, physical and sexual violence in adolescent, heterosexual, intimate relationships. Questionnaires were administered to 487 pupils at two secondary comprehensive schools in Oxford. Data from the questionnaire reveal that a minority of adolescents regularly employ violence in intimate relationships and suggest no significant differences in levels of violence between adolescents of different gender, religious affiliation, household composition or social class. A review of the literature on the use of this standardised questionnaire highlights serious methodological and epistemological problems and questions the use of such questionnaires in future research on the phenomenon of intimate violence.
The primary focus of the thesis concerns transcript data from seventeen single-sex focus groups and thirteen individual interviews. Transcript data reveals that girls and boys recount different experiences of reality. The discourse used by girls and boys represents an active negotiation of personal experience and cultural prescriptions of meaning.
Peers, parents, siblings, teachers, school administrators and media inform adolescents about dominant definitions and boundaries of gender. These definitions are discussed as 'hegemonic masculinity' and 'emphasised femininity' in which gender is structured as distinct, separate, hierarchical and biologically determined. Girls and boys who employed discourses of biological determinism described intimate violence as inevitable and largely a function of female responsibility. Conflict results from the negotiation of this culturally dominant discourse and personal experience. A minority of girls and boys employed other discourses such as those of socialisation and feminism. These discourses provide alternative understandings of personal experience and social identity which some adolescents may find empowering and represent a crucial resistance to the ascendancy of culturally practiced gender.
Donne's Devotions in the context of his early sermons : a revelation of St. John the Divine

This thesis explores the relationship between Donne's Devotions Upon Emergent Occasions (1623) and his early sermons. Its underlying contention is the existence in the Devotions of a highly organized, thematic structure, which not only focusses major concerns scattered through the sermons, but places them in typological relation. Chapters 1-4 concentrate on Donne's discussion of these concerns in the sermons; Chapter 5 then demonstrates their recognizable convergence in the Devotions to trace an evolving narrative.
Chapter 1 begins with beginnings, and Donne's obsessive pursuit of the origin through recreation. This is related to his fascination with the nothingness underlying birth, which he associates with God. His attempts at infinite contraction and evacuation are examined in this context, as are his images of dissolution.
Chapter 2 then turns outwards to Donne's notions of public authority. But in both the political sphere and the Bible, truth is seen to vary with changing aesthetic and political needs, insofar as its ultimate reference points (Biblical translators, King) are themselves less than absolute.
Chapter 3 focusses on the sermons' own attempts therefore to image the absolute, arguing that Donne positions the reader at the forefront of an increasingly refined, textual present through the telescoping of real into narrative time. The sacramental nature of the read word is suggested in this conjunction of temporal passage with liturgical present.
Chapter 4 then examines the idea of division which underlies much of this discussion, as Donne deals with it explicitly and as it is manifested in the sermon structures. The paradox of the Trinity is proposed as a paradigm for Donne's ambivalence; the resolution it allows in terms of union, closure and self-containment is investigated.
Chapter 5, finally, draws these threads of beginning, authority, sacramental reading and division into the tripartite framework of the Devotions, ordering and re-defining them as a series of births that mark the stages of Christian life.
The conjugation system of staphylococcus aureus

A conjugation system in Staphylococcus aureus has been investigated and shown to be determined, at least in part, by genes carried on plasmids. Conjugation required cell-to-cell contact but not calcium ions. The frequency of conjugation depended on the recipient used and on the incubation conditions.
Two conjugative plasmids were mapped by restriction enzyme analysis but experiments to clone the conjugation-determining region were unsuccessful although separate regions specifying gentamicin resistance, ethidium bromide resistance and cadmium resistance were cloned. The gentamicin resistance determinant was probably part of Tn4001. Deletion of various sized pieces of DNA from one of the plasmids resulted in reduction of its ability to specify conjugation but no specific part of this plasmid could be implicated in the process. Further experiments led to the conclusion that this particular plasmid (p8325-4) is probably not self-transmissible but transferred by a phage-mediated system.
Strains of Staphylococcus aureus produced a pheromone-like substance that elicited a clumping response in Streptococcus faecalis but no evidence was found for the involvement of staphylococcal conjugative plasmids in this.
The conjugative plasmid, p8325-2, mobilized a small plasmid (pT181) but not a chromosomal gene.
Insertion of transposon Tn551 was used to produce mutants of the conjugative plasmid p8325-2. Some twenty-six mutants were studied and the position of Tn551 in them mapped. There were preferred regions of insertion for Tn551 and twenty out of the twenty-six mutants had altered ability to conjugate. One showed a significantly higher frequency of conjugation and the other nineteen, all with substantially lower frequencies of conjugation, were mapped to two well-separated regions of the plasmid. Similarity between the locations of these putative regions and those reported for some other conjugative plasmids from staphylococci is striking and suggests a common origin.
Structural studies on manganese superoxide dismutase

Superoxide dismutases are widely distributed enzymes which catalyse the dismutation of superoxide radicals to dioxygen and hydrogen peroxide and are considered to be an important agent of an organism's defence against oxygen toxicity. The crystallization and low resolution structure determination of manganese superoxide dismutase (E.G. 1.15.1.1) from Bacillus stearothermophiluB is described.
The enzyme crystallized in space group P21212 with two monomers per asymmetric unit and cell dimensions of ̲a=72.2Å, ̲b=111.1Å and ̲c=51.1Å. The crystals diffracted to beyond 2Å resolution but were fragile and prone to cell dimension changes. The cell dimension variability was overcome to some extent by crossllnking with glutaraldehyde.
An electron density map was calculated to 6Å resolution initially by the method of multiple isomorphous replacement using data obtained from six heavy atom derivatives. The final map was calculated from single isomorphous replacement data using a map modification procedure. The fitting of an alpha carbon model of iron superoxide dismutase into the map suggested the iron and manganese enzymes are structurally related. The position of the metal atoms in the model solved difference Patterson maps calculated from data collected from a manganese-free crystal and from anomalous dispersion data. The latter data were collected using synchrotron radiation tuned close to the manganese absorption edge. The low resolution map and the availability of 2.4Å resolution native data paves the way for higher resolution X-ray studies of the crystals.
A detailed analysis of amino acid sequences has been carried out on the various metal-containing superoxide dismutases. The results indicate that the enzymes can be classified according to their metal cofactor. The distribution and homology of the enzyme classes supports the endosymbiotic theory of the origin of cell organelles. The presence of the copper/zinc enzyme in Photobacterium leiognathi is shown to support the case for a eukaryote to prokaryote gene transfer.
Carbohydrates in synthesis

The use of carbohydrates as starting materials for organic synthesis is illustrated by the synthesis of several polyhydroxylated piperidine, pyrrolidine, indolizine and pyrrolizidine alkaloids.
Nucleophilic displacement by azide ion at C-2 in a D-glucose derivative, with subsequent intramolecular cyclisation through nitrogen onto the C-6 or C-5 position and functional group manipulation, led to the synthesis of: 1,5-dideoxy-1,5-imino-D-mannitol; 1,2,5-trideoxy-1,5-imino-D-arabinohexitol; 1,5-dideoxy-1,5-imino-D-glucitol; 2-Acetamido-1,5-imino-1,2,5- trideoxy-D-glucitol; 2-Acetamido-1,5-imino-1,2,5-trideoxy-D-mannitol; (2S, 3R, 4R, 5S)-3,4,5-trihydroxypipecolic acid; (2S, 3R, 4R, 5R)-3,4,5- trihydroxypipecolic acid and 2,5-dideoxy-2,5-imino-D-mannitol.
Nucleophilic displacement by azide ion at C-3 in a D-glucose derivative, with subsequent intramolecular cyclisation through nitrogen onto the C-6 position, produced the tosylate salt of 3,6-dideoxy-3,6-imino-1,2-0-isopropylidene- ⫏-D-glucofuranose, a highly divergent intermediate, from which the pyrrolidines: 1,4-dideoxy-1,4-imino-L-gulitol, 1,4-dideoxy-1,4-imino-D-lyxitol and (2S, 3S, 4R) - 3,4-dihydroxyproline were prepared directly. Periodate cleavage of the C1-2 bond and a 2-C chain extension from C-2 with subsequent intramolecular cyclisation, produced the pyrrolizidine (IS, 2R, 8R) - 1,2-dihydroxypyrrolizidine. An intramolecular Wadsworth Emmans cyclisation between a lactol at C-1 and a phosphonate, produced by a DCC coupling of the amine with dimethoxyphosphinylacetic acid, led to the formation of the indolizine (1S, 2R, 8S, 8aR) - 1,2,8-trihydroxyoctahydroindolizine.
The synthesis of 1,4-dideoxy-1,4-imino-D-lyxitol, was achieved by connection of C1 and C4 of a D-mannose derivative through nitrogen.
Methyl 3,5-0-isopropylidene- ⫏-D-xylofuranoside was elaborated to both enantiomers of 1,4-dideoxy-1,4-imino-arabinitol. The _D-enantiomer was produced by introduction of a nitrogen between C-2 and C-5, the L- enantiomer by introduction of a nitrogen between C-1 and C-4.
(2R, 3S, 4R) - 3,4-dihydroxyproline was prepared from _D-ribonolactone. The key step of the synthesis was a nucleophilic displacement by azide ion at C-2 in which the stereochemistry was unexpectedly retained.
The mechanics of continental deformation in Asia and California

The failure of plate tectonic theory to explain the deformation observed across continental plate boundary zones has led to the development of continuum models of lithospheric deformation in which the lithosphere is treated as a layer of viscous material governed by the equation for creeping viscous flow,

ϑσji = ρ9i
ϑxj
where the left hand side represents spatial gradients of stress that are balanced by differences in gravitational potential energy on the right hand side.
Strain rates and an assumed power law rheology are used as a proxy for stress,

τij = BΕ(1/n)-1εij
where B is a strength parameter that represents the depth-averaged temperature dependence of the lithospheric rheology, E is the second invariant of the strain rate tensor, and n is the power law exponent in the stress-strain rate relationship. Finite element methods are used to solve the thin viscous sheet equations (England & McKenzie, 1982), yielding a velocity field that may be compared to observations from regions of continental deformation.
In this study I investigate the deformation of the lithosphere in two regions of active deformation: south-east Asia and the western North America plate boundary zone. Particular attention is paid to the degree to which the deformation fields reflect the driving forces, and the effect of lateral variations in effective viscosity and lithospheric strength upon the deformation field.
For the complete abstract, please consult the pdf.
Clusters of galaxies

The observations on which this thesis are based are of three types:
i) Spectroscopy of southern galaxies from which redshifts have been determined. These redshifts have enabled the calculation of velocity dispersions of the galaxies in five clusters of galaxies: Klemola 44, Sersic 40/6, 2354-35, A496 and the Pavo Group. Redshifts of galaxies in several other southern clusters have also been obtained.
ii) Photographic Photometry using digitised two-dimensional micro-densitometer scans of IIIaJ Schmidt plates of two southern clusters of galaxies: K44 and Sersic 40/6. Similar photometry of two other clusters, A1146 and 2354-35, is also available. Magnitudes in the B-band to a limiting isophote of 27mag/o" are obtained for all galaxies to a limiting magnitude of ~20 within a ~2Mpc (Ho = 50 km s-1 Mpc-1) square around the cluster. Methods of distinguishing stars from galaxies are discussed.
iii) Photoelectric photometry through several circular apertures of 21 bright southern galaxies, using the B, V, R and I bands. As well as providing catalogue data for southern galaxies, these readings supply magnitude zero-points for the photographic photometry of clusters.
The results are discussed in the light of current theories of the morphology, formation and evolution of clusters of galaxies. The X-ray flux from clusters of galaxies is found to be consistent with emission by bremsstrahlung from a hot intracluster gas. Four of the clusters: Klemola 44, 2354-35, A1146 and Sersic 40/6 contain cD galaxies. These galaxies increase in ellipticity with radius and are aligned with their clusters (except A1146); such properties are predicted if cD galaxies evolve by accretion of other galaxies. It is suggested that Klemola 44 may be undergoing a phase of rapid core evolution, leading to the accretion of the core by the cD galaxy. The cluster 2354-35 may have already passed through this evolutionary stage.
Lithostratigraphy and depositional history of the middle triassic dowsing Dolomitic formation of the southern North Sea and adjoining areas

Middle Triassic deposition in Northwest Europe occurred in the enclosed Germanic Basin, which extended from its connection with Tethys in eastern Poland to eastern England.
A detailed lithostratigraphic subdivision of the middle Triassic Dowsing Dolomitic Formation of the U.K. Southern North Sea and subsurface eastern England is presented, based on petrophysical logs, well cuttings and limited core. This subsurface lithostratigraphy is linked to the established outcrop lithostratigraphy of eastern England by means of a surface gamma-ray profile at Kirton Brickworks, Nottinghamshire. The outcrop lithostratigraphy is thereby accurately correlated with that of the basin centre in north Germany.
The major tectonic elements active during middle Triassic deposition in the Southern North Sea area are identified, including previously unreported differential subsidence in an extension of the Dutch Broad Fourteens Basin in the U.K. Sector.
The basinwide Hardegsen unconformity marks the base of the middle Triassic. A transgressive phase resulted first in the deposition of lacustrine pro-delta mudstones towards the basin centre, followed by the deposition of a more extensive thin marine mudstone which formed the base of the middle Triassic over most of the U.K. Southern North Sea area. Two regressive cycles followed. Each started with deposition of the cyclic, hypersaline-marine halites (the Main and Upper Röt Halites), followed by sabkhas, then playa mudflat/lagoonal deposits. At the start of the Muschelkalk deposition, restricted marine conditions were established in the Southern North Sea area. These were followed by a further regressive sequence of hypersaline-marine halites (the Muschelkalk Halite), sabkhas, and finally playa mudstones. The establishment of continental sedimentation across the whole of the Germanic Basin marked the end of the middle Triassic.
The cyclicity within the Germanic Basin correlates with the published coastal onlap charts, indicating its origin is eustatic fluctuations in Tethys.