Protein-tyrosine phosphatases (PTPs) catalyze the hydrolysis of phosphorylated tyrosines by a 2-step mechanism involving nucleophilic attack by cysteine and general acid catalysis by aspartic acid. In most PTPs the aspartic acid resides on a flexible protein loop, consisting of about a dozen residues, called the WPD loop. PTP catalysis rates span several orders of magnitude, and differences in WPD loop dynamics have recently been show to correlate with the rate of enzymatic catalysis. The rate of WPD loop motion could possibly be related to a widely conserved tryptophan residue on the WPD loop. Therefore, point mutants were made in PTP1B (a human PTP) to the conserved tryptophan residue and their effects on catalytic rate and chemical reaction were studied. The results of these studies are presented in this thesis.

<p>In order to extend the applications of aluminium, organic coatings may be applied on sheet materials, for instance for corrosion protection or aesthetic surface finish purposes in the automotive and construction industries, or on foil materials in the flexible packaging industry.</p><p>The most common mechanisms for deterioration and structural failure of organically coated aluminium structures are triggered by exposures to the surrounding environment. Despite the great importance to elucidate the influence of exposure parameters on a buried aluminium/polymer interface, there is still a lack of knowledge regarding the mechanisms that destabilise the structure. It is generally believed that a detailed <i>in situ</i> analysis of the transport of corroding species to the buried interface, or of surface processes occurring therein, is most difficult to perform at relevant climatic and real-time conditions.</p><p>In this work, Attenuated Total Reflection Fourier Transform Infrared Spectroscopy (ATR-FTIR) in the Kretschmann-ATR configuration was successfully applied to <i>in situ</i> studies of the transport of water and ionic species through polymer films to the aluminium/polymer interface upon exposure to ultra pure deionised water and to a 1 M sodium thiocyanate (NaSCN) model electrolyte. Other main processes distinguished were the formation of corrosion products on the aluminium surface and swelling of the surface-near polymer network. Hence, <i>in situ</i> ATR-FTIR was capable to separate deterioration-related processes from each other.</p><p>To perform more unambiguous interpretations, a spectro-electrochemical method was also developed for<i> in situ</i> studies of the buried aluminium/polymer interface by integrating the ATR-FTIR technique with a complementary acting technique, Electrical Impedance Spectroscopy (EIS). While transport of water and electrolyte through the polymer film to the aluminium/polymer interface and subsequent oxidation/corrosion of aluminium could be followed by ATR-FTIR, the protective properties of the polymer as well as of processes at the aluminium surface were simultaneously studied by EIS. The integrated set-up provided complementary information of the aluminium/polymer sample investigated, with ATR-FTIR being sensitive to the surface-near region and EIS being sensitive to the whole system. While oxidation/corrosion and delamination are difficult to distinguish by EIS, oxide formation could be confirmed by ATR-FTIR. Additionally, while delamination and polymer swelling may be difficult to separate with ATR-FTIR, EIS distinguished swelling of the polymer network and also identified ultimate failure as a result of delamination.</p><p>The capability of the integrated ATR-FTIR / EIS <i>in situ</i> technique was explored by studying aluminium/polymer systems of varying characteristics. Differences in water and electrolyte ingress could be monitored, as well as metal corrosion, polymer swelling and delamination.</p>

The main aim of the study in this thesis was to use structure-based protocols to design new drugs for enzymes, DXS and DXR in the non mevalonate pathway. Another aim of this study was to identify the dimer interface in E.coli FabH as an allosteric binding site for designing new class of anti-infective drugs. We have attempted to identify potential inhibitors for DXS by docking the NCI Diversity set compounds, compound libraries available from GSK-MMV and St. Jude's Children's research center. FabH dimer interface has been identified as a potential target using SiteMap, Alanine mutagenesis and docking studies. The first chapter gives an overview of the computational methods. The next two chapters briefly introduce the biological targets in the author's study. Chapter two explains the importance of non-mevalonate pathway in microbes. Different enzymes in the non-mevalonate pathway are discussed and the importance of terpenoids in biological processes and also the use of terpenoids as drugs have been extensively discussed in this chapter. The crystal structures available for DXS and DXR are also discussed. Chapter three brings out the importance of FabH as an anti-infective target. Crystal structure of FabH E.coli is discussed and the importance of FabH as a dimer has been discussed in this chapter. Chapter 3 describes the methods, homology models generated, and analysis from docking studies. The homology models for PvDXS and PvDXR have been used in this study to identify potential inhibitors. Domain swapping and the structural organization of PvDXS before and after domain swaping are discussed. Identification of domain swaping in PvDXS using entropy changes has been extensively discussed. Chapter 4 focuses on FabH (Fatty Acid Biosynthesis, enzyme H also referred to as β-ketoacyl-ACP-synthase III) dimer interface as an allosteric target. SiteMap analysis and MD simulations on the FabH monomer and dimer structures revealed the dimer interface as a binding region. Further analyses were done by mutagenesis studies on the Phe87 residue, a key residue at the dimer interface region and validating the results using docking studies. NCI Diversity Set compounds were docked at the dimer interface of FabH, which revealed that compounds NSC91529 and NSC19803 docked best at the dimer interface region with the phenyl ring of both the compounds

The scope of this work focuses on computationally modeling compounds with protein structures. While the impetus of drug discovery is the innovation of new therapeutic molecules, it also involves distinguishing molecules that would not be an effective drug. This can be achieved by inventing new tools or by refining old tools. Virtual screening (VS, also called docking), the computational modeling of a molecule in a receptor structure, is a staple in predicting a molecule's affinity for an intended target. In our Virtual Target Screening system (also called inverse-docking), VS is used to find high-affinity targets, which can potentially explain absorption, distribution, metabolism, and excretion (ADME) of a molecule of interest in the human body. The next project, low-mode docking (LD), attempts to improve VS by incorporating protein flexibility into traditional docking where a static receptor structure has potential to produce poor results due to incorrectly predicted ligand poses. Finally, VS, performed mostly on small molecules, is scaled up to cyclic peptides by employing Monte Carlo simulations and molecular dynamics to mimic the steps of small molecule VS. The first project discussed is Virtual Target Screening (also called inverse-docking) where a small molecule is virtually screened against a library of protein structures. Predicting receptors to which a synthesized compound may bind would give insights to drug repurposing, metabolism, toxicity, and lead optimization. Our protocol calibrates each protein entry with a diverse set of small molecule structures, the NCI Diversity Set I. Our test set, 20 kinase inhibitors, was predicted to have a high percentage of kinase "hits" among approximately 1500 protein structures. Further, approved drugs within the test set generally had better rates of kinase hits. Next, normal mode analysis (NMA), which can computationally describe the fundamental motions of a receptor structure, is utilized to approach the rigid body bias problem in traditional docking techniques. Traditional docking involves the selection of a static receptor structure for VS; however, protein structures are dynamic. Simulation of the induced fit effect in protein-ligand binding events is modeled by full articulation of the approximated large-scale low-frequency normal modes of vibration, or "low-modes," coupled with the docking of a ligand structure. Low-mode dockings of 40 cyclin dependent 2 (CDK2) inhibitors into 54 low-modes of CDK2 yielded minimum root-mean-square deviation (RMSD) values of 1.82 – 1.20 Å when compared to known coordinate data. The choice of pose is currently limited to docking score, however, with ligand pose RMSD values of 3.87 – 2.07 Å. When compared to corresponding traditional dockings with RMSD values of 5.89 – 2.33 Å, low-mode docking was more accurate. The last discussion involves the rational docking of a cyclic peptide to the murine double minute 2 (MDM2) oncoprotein. The affinity for a cyclic peptide (synthesized by Priyesh Jain, McLaughin Lab, University of South Florida), PJ-8-73, in MDM2 was found to be within an order of magnitude of a cyclic peptide from the Robinson Lab at the University of Zurich in Switzerland. Both are Β-hairpin cyclic peptides with IC50 values of 650 nm and 140 nm, respectively. Using the co-crystalized structure of the Robinson peptide (PDB 2AXI), we modeled the McLaughlin peptide based on an important interaction of the 6-chloro-tryptophan residue of the Robinson peptide occupying the same pocket in MDM2 as the tryptophan residue by the native p53 transactivation helical domain. By preserving this interaction in initial cyclic peptide poses, the resulting pose of PJ-8-73 structure in MDM2 possessed comparable active site residue contacts and surface area. These protocols will aid medical research by using computer technology to reduce cost and time. VTS utilizes a unique structural and statistical calibration to virtually assay thousands of protein structures to predict high affinity binding. Determining unintended protein targets aids in creating more effective drugs. In low-mode docking, the accuracy of virtual screening was increased by including the fundamental motions of proteins. This newfound accuracy can decrease false negative results common in virtual screening. Lastly, docking techniques, usually for small molecules, were applied to larger peptide molecules. These modifications allow for the prediction of peptide therapeutics in protein-protein interaction modulation, a growing interest in medicine. Impactful in their own ways, these procedures contribute to the discovery of drugs, whether they are small molecules or cyclic peptides.

Fast pyrolysis, the rapid thermal decomposition of organic material in the absence of oxygen, is a process that can be used to convert biomass into liquid fuels and chemicals. When performed at the micro-scale, pyrolysis is useful for characterizing biomass structure, as well as determining the pyrolysis products that can be generated from specific biomass feedstocks. Indeed, microscale pyrolysis coupled with on-line analysis of the pyrolysis vapors by GC/MS, so-called pyrolysis-GC/MS (Py-GC/MS), is a technique that can be used to characterize the structure and composition of the various components of lignocellulosic and microalgal biomass based on their pyrolysate distributions. Pyrolysates produced also provide insight into the range of products that can be expected when biomass feedstocks are subjected to thermal decomposition processes. This dissertation focuses on the Py-GC/MS analysis of lignocellulosic biomass such as sorghum and Scenedesmus sp. microalgae, in addition to high-lignin feedstocks such as walnut shells, coconut shells, olive pits and peach pits. The differences in the pyrolysate distributions among these biomass types are correlated with differences in the structure and composition of the biopolymers, mainly cellulose, hemicellulose and lignin, present in the biomass. Py-GC/MS analysis of lignin extracted from endocarp feedstocks is also emphasized. In addition to biomass and extracted lignin, sinapyl (S) and coniferyl (G) alcohol have been analyzed by Py-GC/MS in order to understand the relationship between the corresponding pyrolysates and sinapyl/coniferyl ratios of lignin present in lignocellulosic biomass.

Previous studies from our laboratory and others show a significant increase in levels of both nuclear and mitochondrial DNA and RNA oxidation in vulnerable brain regions in the progression of Alzheimer’s disease (AD). Although total DNA oxidation is increased in AD it remains unclear whether oxidative damage is widespread throughout the genome or is concentrated to specific genes. To test the hypothesis that specific genes are more highly oxidized in the progression of AD, we propose to quantify the percent oxidative damage in genes coding for proteins shown to be altered in the progression of AD using quantitative/real-time polymerase chain reaction (qPCR/ RT-PCR). To further test the hypothesis that diminished DNA repair capacity in the progression of AD contributes to increased DNA oxidation we will use custom PCR arrays and qPCR, Western blot analysis and activity assays to quantify changes in enzymes involved in base excision repair (BER). In order to carry out these studies tissue specimens from superior and middle temporal gyri (SMTG) and inferior parietal lobe (IP), as well as, a non-vulnerable region, the cerebellum (CER) will be analyzed from normal control (NC) subjects and subjects throughout the progression of AD including those with preclinical AD (PCAD), mild cognitive impairment (MCI), and late stage AD (LAD). We will also analyze specimens from diseased control subjects (DC; Frontotemporal dementia (FTD) and dementia with Lewy bodies (DLB)) to determine if the changes we observe in AD are specific.

De flesta klorater som finns i naturen kommer från utsläpp av människan i flera storaindustrier. Kloraterna påverkar flera olika levande organismer och det är därför viktigt att tahand om dem efter utsläppen. Flera olika bakterier kan bryta ned klorat till klorid och syre, enav dessa är Ideonella dechloratans. Nedbrytningen sker med de alkaliska enzymernakloratreduktas och kloritdismutas under anaeroba förhållanden. I detta arbete har 2Dgelelektrofores utvecklats för alkaliska proteiner i Ideonella dechloratans i syfte att kunnajämföra proteinuttryck under olika odlingsförhållanden. Därefter har en jämförelse mellanaeroba och anaeroba odlingsförhållanden gjorts samt försök att detektera kloritdismutas.Resultatet har blivit att en metod har tagits fram som ger en hög grad av upplösning pågelerna från 2D-gelelektroforesen. Jämförelserna mellan aeroba och anaerobaodlingsförhållanden har visat att det finns stora skillnader i proteinuttryck mellan de bådaodlingsförhållandena. Fler proteiner syns i området pH 8-10 under anaerobaodlingsförhållanden jämfört med aeroba förhållanden. Försök till identifiering avkloritdismutas har gjorts för de anaeroba odlingsförhållandena, dock krävs det ytterligarearbete innan en säker identifiering kan göras.

I den reviderade variation av LGR 11 (Läroplan för grundskolan, förskoleklassen ochfritidshemmet 2011) från 2017 lyfts digitaliseringen fram. Skolan är under förändringoch digitala verktyg finns i klassrum. Eleverna förväntas använda dessa och utvecklaförmågor som lyfts fram i LGR 11 och utveckla kunskaper om hur de ska förhålla sigtill ett konstant informationsflöde. Samtidigt använder lärare redan idag många digitalaverktyg i sin undervisning. Med de förändringar som gjorts markerar myndigheter iSverige vikten av att eleverna lär sig navigera och hantera den digitala världen.Frågan som då måste ställas är hur det ser ut i klassrum i Sverige och hurdigitaliseringen har påverkat undervisningen i kemi. När datorn gjorde entréförändrades klassrummet men det tog tid. Idag finns det en mängd olika digitalaverktyg att använda. Genom att gå ut och intervjua lärare skapades en bild avsituationen i det digitala kemiklassrummet. De arbetade med digitala böcker,interaktiva övningar och formativ feedback via delade dokument. Digitaliseringen iskolan speglar den utveckling som samhället har tagit och det är viktigt att alla få följamed i den utvecklingen.Det har även bedrivits forskning inom digitaliseringen i skolan där begrepp som digitalläsning återfinns. Hur har undervisningen hanterats i förhållande till den forskningsom finns och hur lyfter digitaliseringen undervisningen? Genom att göra en jämförelsemellan forskning och praktik skapas en bild av hur det ser ut i skolor men också varmöjligheter för utveckling ligger.

Per-and polyfluoroalkyl substances (PFAS) are anthropogenic compounds that have been spread all over the world. The use of fluorotelomer compounds, short-chained homologues, and other PFASs with perfluorinated moieties has emerged recent years. One of these emerging compound classes is polyfluoroalkyl phosphate esters (PAPs), which have the ability to degrade into persistent PFCAs. The aim of this thesis was to assess the contribution of PAPs and other precursors to the exposure of PFCAs to humans and the environment. The main objective was to analyze a wide range of PFAS in human serum, wild bird eggs, indoor dust, waste water, and sludge. There was a significant contribution from selected precursors to the total amount of PFASs in the abiotic compartments indoor dust, waste water, and sludge. Levels of PAPs found in house dust exceeded those of PFCAs and perfluorosulfonic acids (PFSAs), revealing PAPs as a world-wide important exposure source. A net increase was during waste water treatment was observed for several PFASs in Swedish waste water treatment plants. Together with presence of precursor compounds and intermediates in the influent water and the sludge, this suggest that degradation of PFCA precursors contributed to the increase of PFCAs. Detection of precursors in human serum, together with slow declining trends of PFCAs, revealed an ongoing exposure of PFCAs to the general population of Australia. The diPAPs and the FTSAs were also detected in raptor bird eggs from Sweden from both the terrestrial and the freshwater environment. The precursors concentrations and patterns observed reveal that current regulatory measures are insufficient for the purpose of protecting humans and the environment from PFASs exposure.

The nuclear envelope forms the interface between the nucleus and the cytoplasm. The nuclear envelope consists of the two concentric lipid membranes, the nuclear pores and the nuclear lamina. The inner nuclear membrane contains hundreds of unique transmembrane proteins showing high tissue diversity. Mutations of some proteins in the nuclear envelope give rise to a broad spectrum of diseases called envelopathies or laminopathies. In this thesis, I aimed to study the functional organization of the nuclear envelope by identifying and characterizing interactions between the nuclear envelope proteins. For this, we developed a novel method called the Membrane Protein Crosslink Immuno-Precipitation, which enable identification of protein-protein interactions in the nuclear envelope in live cells. We identified several novel interactions of the inner nuclear membrane protein, Samp1, and studied the interaction between the Samp1 and the nuclear GTPase, Ran in detail. Samp1 can bind to Ran and is thus the first known transmembrane Ran binding protein and Samp1 might provide a local binding site for Ran in the inner nuclear membrane. We found that Samp1 also binds to the inner nuclear membrane protein, Emerin and Ran can regulate the Samp1-Emerin interaction in the nuclear envelope. During mitosis, Samp1 distributes in the mitotic spindle. Therefore, we investigated a possible functional role of Samp1 in the mitotic machinery. Samp1 depletion resulted in aneuploid phenotypes, metaphase prolongation and decreased distribution of γ-tubulin and β-tubulin in the mitotic spindle. We found that Samp1 can bind to γ-tubulin, which is essential for the microtubule nucleation and hence for the spindle stability. The new interesting features of Samp1 provide insights on the unforeseen functions of the nuclear envelope proteins. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 3: Manuscript. Paper 4: Manuscript.</p>