Les systèmes neuronaux et vasculaires nécessitent un réseau complexe pour exécuter correctement leurs fonctions. Les processus impliqués dans la création de ce réseau s'appuient sur des voies coordonnées, souvent activées par des systèmes protéine/récepteur communs, qui conduisent au remodelage du cytosquelette.En général, les cellules neuronales et vasculaires répondent aux stimuli extracellulaires sous forme de protéines solubles sécrétées, qui interagissent avec les récepteurs de surface pour favoriser l'attraction ou la répulsion vers la source des protéines sécrétées. Ce processus, appelé guidage, est régulé par sept familles de récepteurs et leurs ligands respectifs, qui s'influencent les uns sur les autres et peuvent agir sur le système neuronal, le système vasculaire ou les deux ensembles.Cette étude est centrée sur deux récepteurs transmembranaires à passage unique, les membres des familles de protéines Roundabout et UNC5 qui sont principalement impliquées dans l'angiogenèse: Robo4 et UNC5B.L'information structurelle sur la région extracellulaire de plusieurs de ces récepteurs, et comment le signal est relayé à travers la membrane, fait défaut.La déglycosylation enzymatique a confirmé que les domaines extracellulaires de Robo4 et UNC5B sont largement glycosylés avec des glycanes liés en azote du complexe type. La mutagenèse dirigée des sites de glycosylation prédits de Robo4 perturbe son expression, indiquant que ces résidus sont nécessaires pour la stabilité de la protéine et que leur glycosylation, ou leur passage dans la voie de glycosylation, pourrait être nécessaire pour un repliement correct. Les données MALS et SAXS montrent qu'en solution, Robo4 ecto est un monomère flexible de forme allongée. Les domaines ne présentent pas des caractéristiques distinctes dans le modèle construit à partir des données SAXS. Plusieurs Fabs se liant au domaine extracellulaire de Robo4 ont été caractérisés, avec l'espoir d'identifier les Fab qui pourraient inhiber l'interaction Robo4 / UNC5B rapportée pour une caractérisation plus poussée. La formation du complexe a été vérifiée par SEC-MALS et SAXS, et les constantes d'interaction ont été déterminées en utilisant SPR. Des cristaux de certains complexes domaine Fab / domaine extracellulaire Robo4 ont été produits, bien que la structure du complexe n'ait pas pu être résolue à l'heure actuelle.Les expériences de pull-down, SEC-MALS et SPR montrent que Robo4 ecto et UNC5B ecto n'interagissent pas entre elles, malgré une étude par un autre groupe montrant le contraire. Étant donné que différentes lignées cellulaires ont été utilisées, des modèles de glycosylation spécifiques, ou une tierce partie non détectée, pourraient être nécessaires pour l'interaction. En raison de leur implication avec les récepteurs extracellulaires, les héparanes sulfates sont un candidat probable, mais d'autres partenaires devraient être envisagés.La structure cristallographique de l'UNC5B ecto est similaire aux structures existantes de UNC5A et UNC5D. Le haut degré de conservation des domaines del’Ig pourrait être une indication de l'importance de cette région, qui est responsable de la liaison à Netrin. Bien que la région de liaison de Netrin soit connue pour être dans les domaines Ig, le site de liaison précis n'a pas encore été déterminé, mais il pourrait être situé à proximité ou à l'intérieur des surfaces chargées négativement présentes sur les domaines Ig observées dans la structure d’UNC5B.Le travail présenté ici devrait servir de base à une meilleure caractérisation biochimique et structurale des récepteurs extracellulaires Robo4 et UNC5B. / Neuronal and vascular systems require a complex network to properly perform their functions. The processes involved in creating this network rely on coordinated pathways, often activated through common protein/receptor systems, which lead to cytoskeletal remodelling. In general, neuronal and vascular cells respond to extracellular stimuli in the form of soluble secreted proteins, which interact with surface receptors to mediate attraction or repulsion towards the source of the secreted proteins. This process, called guidance, is regulated by seven families of receptors and their respective ligands, which influence each other and can act on the neuronal system, the vascular system or both.Structural information about the extracellular region of many of these receptors, and how signal is relayed across the membrane, is lacking.This study is focused around the extracellular domain of two single-pass transmembrane receptors of the Roundabout and UNC5 protein families that are majorly involved in angiogenesis: Robo4 and UNC5B.Based on the findings of this study, the Robo4 and UNC5B extracellular domains are extensively glycosylated with N-linked glycans of the complex type. Site-directed mutagenesis of the predicted Robo4 glycosylation sites disrupts protein expression, indicating that they are necessary for protein stability and passage through the glycosylation pathway might be necessary for correct folding. MALS and SAXS data show that in solution the Robo4 extracellular domain is a flexible monomer with extended shape. Several Fabs binding to the extracellular domain of Robo4 were characterised, with the expectation to identify those Fabs that could inhibit the reported Robo4/UNC5B interaction for further characterisation. Complex formation was verified by SEC-MALS and SAXS, and interaction constants were determined using SPR. Crystals of some Robo4 extracellular domain/Fab complexes were produced, although the structure of the complex could not be solved at the present time.Despite a study by another group showing otherwise, pull-down, SEC-MALS and SPR experiments show that the Robo4 and UNC5B extracellular domains do not interact with each other. It is proposed that the difference may be caused by different glycosylation patterns specific to the cell lines used for each study, or by an undetected third party necessary for interaction. This, however, still requires further study. SEC-MALS analysis showed that the UNC5B extracellular domain is a monomer in solution and its crystal structure was solved at 3.4 Å resolution. Comparison to the existing structures of human UNC5A and rat UNC5D shows striking similarities and a high degree of evolutionary conservation of the Ig domains might be indication of the importance of this region, which is responsible for binding to the guidance cue Netrin. Although the Netrin binding region is known to be within the Ig domains, the precise binding site has not yet been determined, but it might be located in proximity, or within, the negatively charged surfaces present on the Ig domains which are observed in the UNC5B structure.It is hoped that the work presented here will give the basis for better biochemical and structural characterisation of these two receptors.

京都大学 / 0048 / 新制・課程博士 / 博士(理学) / 甲第19497号 / 理博第4157号 / 新制||理||1597(附属図書館) / 32533 / 京都大学大学院理学研究科物理学・宇宙物理学専攻 / (主査)教授 鶴 剛, 教授 谷森 達, 准教授 成木 恵 / 学位規則第4条第1項該当 / Doctor of Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(理学) / 甲第20912号 / 理博第4364号 / 新制||理||1626(附属図書館) / 京都大学大学院理学研究科物理学・宇宙物理学専攻 / (主査)教授 井岡 邦仁, 教授 川合 光, 教授 田中 貴浩 / 学位規則第4条第1項該当 / Doctor of Science / Kyoto University / DGAM

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第21765号 / 工博第4582号 / 新制||工||1714(附属図書館) / 京都大学大学院工学研究科材料工学専攻 / (主査)教授 河合 潤, 教授 神野 郁夫, 准教授 奥田 浩司 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20071号 / 医博第4164号 / 新制||医||1018(附属図書館) / 33187 / 京都大学大学院医学研究科医学専攻 / (主査)教授 黒田 知宏, 教授 松田 文彦, 教授 楠見 明弘 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

The science and technology of X-rays has only been part of human achievement for the past 100 years, while the study of image formation in general has endured for as long as 1000 years. The ability to conceive, design, and fabricate X-ray imagers, moreover, has existed for only the past 70 years, and X-ray astronomical telescopes have been in use for a mere 35 years. Considering that aplanatic, normal incidence telescope designs required more than 400 years to perfect, it is most interesting to note that the development of ‘aplanatic’ grazing incidence telescopes has taken only about 40 years. In order to improve and expand the field of X-ray astronomy, and imaging in general, we find that these days a comprehensive systems engineering approach to X-ray image formation must be undertaken. While some industrial interests have taken steps in this direction, any academic approach is lacking from within the archival literature to date, and there are virtually no established university courses. Indeed, it would seem that top level, optical-systems-engineering is exclusively reserved for those seasoned professionals who have accumulated (though somewhat artistically) the “know-how” to efficiently conceive and implement excellent optical designs. Such expert knowledge is not and should not be mysterious. To this end, we attempt to formulate a highly comprehensive approach to X-ray optical systems engineering and implement it within the context of the Wolter Type-I and Type-II (grazing incidence) telescopes currently utilized for practical X-ray/EUV astronomy. In addition, we will transform the classical paraboloid-hyperboloid designs into ‘aplanatic’ and ‘isoplanatic’, hyperboloid-hyperboloid systems, where certain coma conditions are minimized. As will be shown, one gains little improvement in performance when choosing a quasi-aplanatic mirror design over a classical one, owing to scatter and other image degradation effects. Next we will show that a generalized hyperboloid-hyperboloid design can be comprehensively optimized for any imaging requirement, where the operational field-of-view is weighted according to spatial information content. Our H-H design has been optimized for the GOES Solar X-ray Imager mission and adopted by NASA and NOAA. It is currently undergoing fabrication by Raytheon Optical Systems Inc. who is under subcontract to the Lockheed-Martin Solar and Astrophysics Laboratory. Our design is expected to result in an 80% increase in optical system performance over the original SXI baseline design.

<p>Arsenic (As) is an element that is well known for its toxic capabilities. It is odorless and colorless and is known to contaminate the drinking water of populations in several parts of the world. Routine monitoring of arsenic exposure is usually performed with urine, hair or nail, where samples are collected for laboratory analysis. Arsenic’s strong affinity to keratin rich tissues make skin another possible measurement site, in addition to the latter two tissues mentioned above. In some cases, skin samples are extracted for analysis. This is painful and invasive and is not ideal for <em>in vivo</em> monitoring of arsenic. The ability to quantify elemental concentration non-destructively is the major calling card of x-ray fluorescence (XRF). To that end, work was started on development of XRF detection systems for arsenic. The technique has shown promise for other elements and dramatic improvements in As detection capabilities were previously found when going from a radioisotope-based x-ray source to an x-ray tube based approach.</p> <p>This thesis documents the comparison of three x-ray tube based detection systems intended for the measurement of arsenic in skin. Two benchtop systems were used, with a) extended development of the previously assembled system and b) the first use of a separate detection system. Two handheld x-ray analyzers (portable detection systems) were also investigated in stand mode, where they were attached to a purpose-built mounting stand, provided by the manufacturer, during all analysis. Polyester resin phantoms were used to model arsenic in skin and a nylon backing was used to represent as bulk tissue behind skin. During the course of the work, modifications were made to the laboratory setup associated with the benchtop approaches.</p> <p>A benchtop polychromatic Mo anode x-ray tube based x-ray fluorescence (XRF) detection system was the first system used in this work. Through modifications to the existing design of the system, the lowest minimum detection limit (MDL) achievable was found to be (0.611±0.001) ppm normalized to gross scatter, where ppm is ug of arsenic per gram of dry weight (resin). The measurement time was ~1800 seconds real time. The equivalent (skin) and whole body effective doses delivered were (19±3) uSv and (163±47) uSv respectively. The corresponding direct (un-normalized) MDL was (0.499±0.002) ppm, in agreement with that found previously. Modifications to the system allowed a reduction in the localized effective dose delivered, to achieve this MDL, from (0.64±0.03) uSv previously to (0.14±0.04) uSv here.</p> <p>Next, the current work investigated two handheld x-ray analyzers provided by InnovX. A PiN diode detector based Alpha 4000S model unit (W anode x-ray tube) and a Silicon Drift Detector (SDD) based Delta model (Au anode x-ray tube). Both units were operated in benchtop mode: they were mounted in a stand and a phantom was placed on a kapton exit window. The lowest gross-scatter normalized and direct detection limit with the Alpha 4000S unit was (1.649±0.002) ppm and (1.651±0.002) ppm respectively. The equivalent and whole body effective doses delivered were found to be (9.4±2.2) mSv and (94±22) uSv respectively. The localized effective dose was (6.4±1.5) X 10^-3 uSv. By comparison, the Delta unit produced a gross-scatter and direct normalized detection limit of (0.570±0.002) ppm and (0.558±0.002) ppm respectively. The equivalent dose delivered was found to be (19.0±9.0) mSv. The corresponding localized and whole body effective doses delivered were (9.7±4.6) X 10^-3 uSv and (190±90) uSv respectively.</p> <p>The last system used in the current research was a monochromatic Ag anode x-ray tube based XRF setup. A doubly curved crystal (DCC) was used to select the Ag K-alpha line and focused the beam to a spot size of mm² at the focal length. The phantoms were placed at a farther distance where the beam had expanded to a larger area. The lowest Compton scatter normalized detection limit with the Si(Li) detector was found to be (0.696±0.002) ppm. After characterizing its performance in a range of energies, a silicon drift detector was also used on this system. It had the benefit of higher throughput capabilities and superior resolution. The housing of the detector was sufficiently small that it could be placed closer to the phantom surface than the Si(Li) detector. The lowest Compton-scatter normalized detection limit with the SDD was (0.441±0.003) ppm in 1800 seconds real time. The equivalent dose was found to be (11±2) mSv and the localized and whole body effective doses were found to be (3.92±0.87) X 10^-3 uSv and (110±23) uSv respectively. A significantly lower system dead time was observed with the SDD. Finally, Monte Carlo simulations of the system were performed to evaluate the performance of three ratios when their phantom measurement values were compared against simulations of skin. Results were found to be in agreement to withinin vivo concentration of arsenic in skin (ICRP).</p> <p>Finally, EDXRF measurements were performed on bulk cores of skin, <em>ex vivo</em>. While it was not possible to detect arsenic in the samples, due to the samples being collected from members of the public as opposed to an exposed population, a depth profile of numerous skin samples, starting from the surface and running straight down, was obtained for calcium, iron and copper.</p> / Doctor of Philosophy (PhD)

All living organisms must organize their genome so that it not only fits within the cell, but remains accessible for cellular processes. In bacteria, an arsenal of nucleoid-associated proteins contributes to chromosome condensation. A novel nucleoid-associated protein was recently discovered in actinobacteria, and is essential in Mycobacterium. It was classified as an integration host factor protein (IHF); however, it does not share sequence or structural homology with the well characterized Escherichia coli IHF. In this study, we characterize the structure and function of Streptomyces coelicolor IHF (sIHF). We have used a combination of biochemistry and structural biology to characterize the role of sIHF in DNA binding and DNA topology. We have solved crystal structures of sIHF bound to various double-stranded DNA substrates, and show that sIHF is able to contact DNA at multiple surfaces. Furthermore, sIHF inhibits the activity of TopA, impacting DNA topology in vitro. Our work demonstrates that sIHF is a novel nucleoid-associated protein with key roles in condensing DNA. We believe that sIHF performs its function by differentially using multiple nucleic-acid binding surfaces. Further characterization is required to confirm this hypothesis in vivo. Given that the Mycobacterium homolog of sIHF (mIHF) is essential, our studies lay the foundation to explore novel drug targets for Mycobacterium tuberculosis and Mycobacterium leprae. / Thesis / Master of Science (MSc) / Unconstrained, the bacterial genome exceeds the size of the cell by 1 000- 10 000 times; thus, compacting it into a condensed structure, known as the nucleoid, is essential for life. An arsenal of nucleoid-associated proteins contributes to this process. In this study, we characterize the structure and function of a novel nucleoid–associated protein from the soil dwelling organism Streptomyces coelicolor. We used a combination of genetics, biochemistry, and structural biology to characterize the role of this protein in DNA binding and nucleoid organization. Since this protein is also found in important human pathogens, this work lays the foundation to explore the use of nucleoid-associated proteins as antimicrobial drug targets.

<p>Metalloproteins, or proteins with a metal ion cofactor, are essential for biological function of both lower and higher level organisms. These proteins provide a multitude of functions from molecular transport, such as the hemoglobin transport of oxygen, to biologically important catalytic processes. As an example case, photosystem II (PSII) is studied as a representative metalloprotein. It was chosen based on the potential impact in the energy sector due to its ability to perform water oxidation using solar based energy. Understanding mechanisms by which the Mn₄Ca cluster inside PSII, also known as the oxygen evolving complex (OEC), can store energy as redox equivalents for splitting water will be essential for future development of analogous artificial systems. By using time resolved x-ray spectroscopy, the electron structure of the metal in the protein was probed through the catalytic cycle. While the applications mentioned herein are based on PSII from spinach, the developments in time-resolved x-ray spectroscopy techniques are also applicable to other metalloproteins.</p><p></p><p>By creating a new x-ray spectrometer we were able to capture the difference in x-ray emission spectra between two compounds differing in a single metal bound ligand, i.e. Mn^IV-OH and Mn^IV=O. This both establishes the functionality of the x-ray emission spectrometer and provides useful insight into the expected changes upon an oxygen double bond formation. This change in spectroscopic signal is discussed in context of the OEC which has been hypothesized to form a Mn^IV=O state.</p><p></p><p>A new sample delivery system and further developments to the x-ray spectrometer enabled both time-resolved x-ray absorption and time-resolved x-ray emission of PSII. These experiments show the potential of synchrotron sources for time-resolved x-ray spectroscopy. From our x-ray absorption measurements we were able to follow the electronic structure changes in time using a single incident photon energy. From the kinetic traces obtained, we show possible alternative interpretations of previous results showing a delay in reduction during the final step in water oxidation. From the x-ray emission spectroscopy (XES) measurements of PSII we were able to reproduce previous results within a limited collection time and give estimates for data size requirements for metalloproteins using this spectrometer. Between the results of both these measurements, we show the improved capability for time resolved measurements at synchrotrons.</p><p>The development of x-ray free electron lasers (XFELs) has also opened many opportunities for understanding faster electronic dynamics by providing femtosecond x-ray pulse durations with ~10¹² photons per pulse. While theoretical modeling of distortions to crystallographic data have been performed, little to no work has been done to understand under what conditions such an intense pulse will have on an impact on emission spectra. Here an atomistic model was developed, and data collected, to clarify the effects of sequential ionization, i.e. two single photons absorbed by the same atom at different times during a single pulse. Experimentally we found that XFELs easily achieve flux densities that invoke a different response than is classically observed for single photon absorption and emission for Mn^II which was used as a representative case for 3d transition metals in general. We also give parameters by which the onset of this damage can be predicted and an approximation to its effect on 3d transition metals. Additionally this work guides the work of future XFEL facilities as it shows that shorter pulses, currently believed to be able to escape x-ray induced distortions to crystallography data, is not a viable method for overcoming changes in x-ray emission spectra.</p><div><br></div>

The exploitation of the soft x-ray/extreme ultra-violet (EUV) region of the electromagnetic spectrum is possible mainly due to the development of multilayer (ML) mirrors. This region of the electromagnetic spectrum offers great opportunities in both science and technology. The shorter wavelength allows one to see smaller features in microscopy and write finer features in lithography. High reflectivity with moderate spectral bandwidth at normal/near-normal incidence can be achieved in soft x-ray/ EUV spectral range using these ML mirrors, where natural crystals with the required large periodicity are not available. These MLs are generally artificial Bragg’s reflectors, which consist of alternative high and low density materials with periodicity in the nanometer range. The main advantages of ML optics stem from the tunability of layer thickness, composition, lateral gradient, and the gradient along the normal to the substrate; these can be tailored according to the desired wavelength regime. They have the great advantage of being adaptable to figured surfaces, enabling their use as reflective optics in these spectral regions, for focusing and imaging applications. Broadband reflectivity and wavelength tunability are also possible by using MLs with normal and lateral gradient, respectively. However, fabrication of these ML mirrors requires the capability to deposit uniform, ultra-thin (a few angstroms-thick) films of different materials with thickness control on the atomic scale. Thus, one requires a proper understanding of substrate surfaces, individual layers, chemical reactivity at interfaces and, finally, of the ML structures required for particular applications. The performance of these MLs is limited by (the lack of) contrast in optical constants of the two materials, interfacial roughness, the chemical reactivity of two materials and, finally, errors in the thickness of individual layers. Soft x-ray/extreme ultra-violet ML mirrors have found a wide range of applications in synchrotron radiation beam lines, materials science, astronomy, x-ray microscopy, x-ray laser, x-ray lithography, polarizers, and plasma diagnostics. The Indus–1 synchrotron radiation (SR) source is an operational 450 MeV machine, which produces radiation up to soft x-rays. Indus-2 is a 2.5 GeV machine, which has been commissioned recently to produce hard x-rays (E > 25 keV). The combination of Indus-1 and Indus-2 will cover a broad energy spectrum from IR to hard x-rays. Therefore, there is a significant need and opportunity to study MLs of different pairs of materials, with different parameters such as periodicity and optimum thickness of individual layers. The goal of the present thesis is to fabricate MLs for soft x-ray optics and to study their physics for application as polarizers in the wavelength range from 67 Å to 160 Å on the Indus-1 synchrotron source. To accomplish this task, a UHV electron beam evaporation system has been developed indigenously for the fabrication of MLs. Three different ML systems viz., Mo/Si, Fe/B4C and Mo/Y have been fabricated, and their surfaces and interfaces were investigated thoroughly for the polarizer application. X-ray reflectivity (XRR) has been used extensively in the investigations of these MLs. This is because XRR is a highly sensitive non-destructive technique for the characterization of buried interfaces, and gives microscopic information (at atomic resolution) over a macroscopic length scale (a few microns). Numerical analysis of XRR data has been carried out using computer programs. Depth-graded x-ray photoelectron spectroscopy (XPS) has been used for compositional analysis at interfaces for some of the ML structures, as a technique complementary to XRR. The performance of some of these MLs has been tested in the soft x-ray region, using the Indus-1 synchrotron radiation (SR) source. Prior to studying the MLs, a detailed study of the surfaces and interfaces of thin films, bi-layers, and tri-layers was carried out using XRR and the glancing incidence fluorescence technique. The discontinuous-to-continuous transition and the mode of film growth, which are vital to the optimization of layer thickness (basically for the high-atomic number or high-Z layer) in the ML structures, were also investigated using in situ sheet resistance measurement method. Indus-1 is a soft x-ray SR source that covers atomic absorption edges of many low-Z materials. The present work demonstrates the possibilities of characterizing low-Z thin films and multilayers using soft x-ray resonant reflectivity. In one case, we have shown for first time that soft x-ray resonant reflectivity can be employed as a non-destructive technique for the determination of interlayer composition. In a second study using the Indus-1 SR source, we have shown, by observing the effect of the anomalous optical constant on reflectivity pattern when photon energy is tuned across the atomic absorption edge of the constituent low-Z element, that soft x-ray resonant reflectivity is an element-specific technique. This thesis is organized into 7 chapters. A brief summary of individual chapters is presented below. Chapter 1 gives a brief general introduction to x-ray ML optics. This is followed by a discussion of the importance of the soft x-ray region of electromagnetic radiation. The optical properties of x-rays are reviewed and optical constants are calculated for some of the important materials used for x-ray MLs. The refractive index in the x-ray region being less than unity (except absorption edges), the consequent limitation of conventional transmission lenses is discussed. The limitation of glancing angle incidence optics is presented, motivating the need for ML optics, which is discussed along with a theoretically calculated reflectivity profile. The procedure for materials for the MLs for application in different spectral regions is discussed, along with a survey of literature related to the present thesis. The importance of the quality of surfaces and interfaces on the performance of ML structures has been shown through simulations. The applications of soft x-ray MLs are discussed with emphasis on polarization. This is followed by a review of different modes of growth of thin films. Finally, the scope of the present work is highlighted. Chapter 2 provides brief descriptions of the experimental techniques used in the present investigations and of the numerical methods employed for quantitative data analysis. The XRR technique is discussed elaborately because it has been used extensively. Detailed calculations of x-ray reflectivity from single surfaces, thin films and bi-layers are presented, along with simulated values. The effect of critical angle and Brewster’s angle is also discussed. Data analysis methods for computing x-ray reflectivity from multilayer structures, based on dynamical and kinematical models, have been discussed. The effect of roughness on XRR has been discussed based on the recursion formalism of dynamical theory. Simulations of XRR and experimental XRR data fitting are carried out using computer programs. The XRR experimental set up is also outlined. A theoretical background is given for the electrical measurements on thin films. This is followed by a brief overview of x-ray photoelectron spectroscopy (XPS) and interpretation of spectra. Finally, the glancing incidence x-ray fluorescence (GIXRF) technique is outlined. Chapter 3 describes in detail the ultra-high vacuum electron beam evaporation system developed in house especially for the fabrication of thin films and x-ray multilayer optics. At the outset, a brief overview of different deposition techniques commonly used for the fabrication of x-ray optical elements is presented. Design, fabrication, and assembly of different accessories are discussed. The control of thickness and uniformity of the films deposited has been checked through the experiments, whose results are provided. The results obtained for ML test structures are presented to show the capability of system in carrying out fabrication of high quality x-ray ML structures. Finally, the versatility of evaporation system incorporating in situ characterization facilities such as -situ electrical measurements for different substrate temperatures is illustrated. Chapter 4 presents a study of the growth of ultra-thin Mo films at different substrate temperatures using in situ sheet resistance measurements. First, a theoretical background is given on the different stages of island growth and on factors affecting thin film growth, followed by a discussion of the possible electrical conduction phenomena in continuous and discontinuous metal films. The nature of thin film growth and a detailed microscopic picture at different growth stages are derived from a modeling of sheet resistance data obtained in situ. The various conduction mechanisms have been identified in different stages of growth. In the island growth stage, the isotropic and anisotropic growth of Mo islands is identified from the model. In the insulator-metal transition region, experimentally determined values of critical exponent of conductivity agrees well with theoretically predicted values for a two-dimensional (2D) percolating system, revealing that Mo films on float glass substrate is predominantly a 2D structure. The minimum thickness for which Mo films becomes continuous is obtained as 1.8 nm and 2.2 nm for Mo deposited at substrate temperatures 300 K and 100 K, respectively. An amorphous-to- crystalline transition is also observed, and discussed. Chapter 5 covers the detailed study of the surfaces and interfaces studies in three different ML structures viz., Mo/Si, Fe/B4C and Mo/Y, meant for the polarizer application in the wavelength range of 67 Å to 160 Å. Multilayers with varying periodicity, varying number of layer pairs, and different ratios of high-Z layer thickness to the period, were fabricated using the electron beam system. Initially, a brief overview of the design aspects of ML structures is given, along with the theoretically calculated reflectivity at Brewster’s angle from the best material combinations. In Mo/Si MLs, the interlayer formed at the interfaces due to interdiffusion of the two elements is asymmetric in thickness, i.e., Mo-on-Si interlayer is thicker than the Si-on-Mo interlayer. To take account of these interlayers in XRR data fitting, a four layer model is considered. The effect of interlayers on reflectivity pattern was studied using simulations, and differences with respect to roughness are also discussed. The mechanism of formation of asymmetric interlayers is also discussed. The interlayer composition has determined using depth-graded XPS. The results reveal the formation of the MoSi2 composition at both the interfaces. The experimental results agree well with theoretical calculations based on solid-state amorphization reaction, which is a result of large heat of mixing. The effective heat of formation model reveals the formation of MoSi2 as the first phase. The soft x-ray reflectivity performance of the Mo/Si ML structure at Brewster’s angle is tested using Indus-1 synchrotron radiation (SR). Using XRR and GIXFR, a study of the surfaces and interfaces of bilayers of B4C-on-Fe and Fe-on- B4C, and tri-layers of Fe-B4C-Fe was carried out, with a systematic variation of Fe and B4C layer thicknesses. A sharp interface was observed in Fe-on-B4C, whereas a low density (w.r.t. Fe) interlayer is observed at the B4C-on-Fe interface. The interlayer properties fluctuates w.r.t. the bottom Fe layer thickness and is independent of the top B4C layer thickness. The nature of fluctuations has been discussed in detail. A study of the surfaces and interfaces of Fe/B4C MLs is described. Finally, a study of the surfaces and interfaces of bilayers, tri-layers, and MLs of the Mo/Y system is discussed in detail. Chapter 6 describes the application of soft x-ray resonant reflectivity for the characterization of low-Z thin films and interfaces in multilayer structures. Initially, a discussion of the energy dependence of atomic scattering factors and hence of optical constants is provided with simulations, with emphasis on the atomic absorption edge. Then, a brief overview of synchrotron radiation, with particular emphasis on the parameters of the Indus-1 synchrotron source is given. The possibilities of determining the composition of the buried interlayer with sub-nanometer scale sensitivity using soft x-ray resonant reflectivity are discussed. The methodology has been applied to study the Mo/Si interface both by simulations and by experiments on the Indus-1 SR, by tuning the photon energy to the Si L-absorption edge. Finally, direct evidence of elemental specificity of soft x-ray resonant reflectivity through the observation of the effect of anomalous optical constants on the reflectivity pattern is discussed. We demonstrate the method through simulations and experiments on the B4C material in B4C thin films and Fe/ B4C bi-layers, using Indus-1 SR tuned to the boron Kedge. Chapter 7 summarizes the main findings of the present work, and provides an outlook for further investigations in the field.