L'émergence des périphériques de capture numériques ont permis le dé- veloppement de l'acquisition 3D pour numériser les propriétés d'un objet réel : sa forme et son apparence. Ce processus fournit une représentation dense et précise d'objets réels et permet de s'abstraire d'un processus de simulation physique coûteux pour modéliser un objet. Ainsi, les problématiques ont évolué et portent non plus uniquement sur la modélisation des caractéristiques d'un objet réel mais sur les traitements de données issues de l'acquisition pour intégrer une copie de la réalité dans un processus de synthèse d'images. Dans ces travaux de thèse, nous proposons de nouvelles représentations pour des fonctions d'apparence issues de l'acquisition dont le but est de dénir un ensemble de modèles multi-échelles, de faible complexité en taille, capable d'être visualisé en temps réel sur le matériel graphique actuel.

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En aquest treball de tesi doctoral es presenta l’estudi del magnetisme de diferents materials de naturalesa metal•lorgànica emmarcats en diferents famílies. Per un cantó, tenim molècules d’alt espí amb anisotropia magnètica uniaxial negativa. Aquesta anisotropia genera una barrera d’energia en aquestes molècules que impedeix al moment magnètic girar lliurement i l’orienta de forma preferent en una direcció de l’espai. El moment magnètic pot canviar el seu sentit d’orientació superant la barrera d’energia de forma clàssica per activació tèrmica o quànticament per efecte túnel. Aquesta propietat es pot utilitzar per aplicacions tecnològiques d’aquestes molècules en el camp de la gravació magnètica d’ultra-alta densitat o en el camp de la computació quàntica, com a qubits magnètics. En la primera part d’aquesta tesi es presenten els estudis de les propietats magnètiques d’algunes d’aquestes molècules pertanyents a la família del Mn12, en diferents entorns (des de monocristalls fins a partícules polimèriques passant per dissolucions), així com noves tècniques de caracterització magnètica d’aquestes molècules com és el dicroisme circular magnètic. Aquest mètode ens permet per primera vegada mesurar alguns fenòmens de física fonamental en aquestes molècules mitjançant caracterització magneto-òptica, com és ara l’efecte túnel del moment magnètic macroscòpic. En el segon bloc d’aquesta tesis es presenta l’estudi del magnetisme d’una nova família de materials nanoporosos altament innovadors, sintetitzats a partir de radicals trifenilmetílics perclorats (PTM), funcionalitzats amb diferents grups carboxílics. L’ús d’aquests radicals ens ha portat a l’obtenció no només de materials nanoporosos amb un alt grau de porositat i unes propietats magnètiques destacables. La cristal•lització d’aquests radicals ha permès la obtenció del primer ferromagnet nanoporós purament orgànic, amb una temperatura de transició del ordre de 125 mK. La cristal•lització de combinacions d’aquests radicals amb ions de metalls de transició en la aproximació metal-orgànica ha donat lloc a varis sistemes notables. La combinació de ions de Cu(III) amb radicals PTM ens permet obtenir la primera esponja magnètica, material nanoporós amb uns 3 nm de diàmetre i amb propietats de pèrdua i reabsorció de solvents reversibles i detectables a través de les seves propietats magnètiques, convertint-se per tant en un sensor magnètic selectiu a certs dissolvents. Per altra banda, la combinació de ions de Co(II) amb els radicals PTM ens porta a la primera estructura nanoporosa magnètica formada per sistemes helicoïdals no inter penetrats i que presenta uns fenòmens de relaxació lenta molt inusuals: aquesta relaxació presenta una forta dependència amb el camp magnètic aplicat y pot ser descrita mitjançant el model de Davidson-Cole. Així doncs, en aquesta tesi es presenten l’estudi de les propietats magnètiques de diversos materials diferents les propietats dels quals poden ser aprofitades sens dubte per aplicacions d’alt nivell tecnològic que van des de la computació quàntica a sensors magnètics de baixa temperatura, passant per la gravació magnètica d’alta densitat. Els resultats presentats han donat lloc a una sèrie de publicacions en revistes científiques d’alt nivell que es detallen a continuació: 1. Gerbier Ph., et al., Synthesis and Characterization of a [Mn12O12(O2CR)16(H2O)4] Complex Bearing Paramagnetic Carboxylate Ligands. Use of a Modified Acid Replacement Synthetic Approach, Monatshefte für Chemie 134 (2003) 265-276 2. Maspoch D.,et al., A nanoporous molecular magnet with a reversible solvent-induced mechanical and magnètic properties, Nature Materials, 2 (2003) 190-195 3. Ruiz-Molina D.,et al., Isolated Single-Molecule Magnets on the Surface of a Polymeric Thin Film, Advanced Materials, 15 (2003) 42-45 4. Domingo N., et al., Synthesis and Characterization of a new chiral nanomagnet, Polyhedron, 22 (2003) 2355-2358 5. Domingo N., et al., Magnetism of Isolated Mn12 Single-molecule Magnets Detected by Magnetic Circular Dichroism: Observation of Spin Tunneling with a Magneto-optical Technique, Physical Review B 69 (2004) 052405 6. Maspoch D., et al., Self-assembly of a Dicarboxylic Radical: a New Pure Organic Robust Paramagnetic Nanoporous Molecular Material, Journal of the American Chemical Society, 126 (2004) 730-731 7. Maspoch D., et al., A New Robust Nanoporous Pure Organic Magnet, Angewandte Chemie Int. Ed. 43 (2004) 1828-1832 8. Gerbier Ph., et al., Chiral, single-molecule nanomagnets: synthesis, magnètic characterization and natural and magnètic circular dichroism, Journal of Materials Chemistry 14 (2004) 2455-2460 9. Maspoch D., et al.,Open-shell nanoporous salts formed by the supramolecular assembly of a polycarboxylate perchlorinated triphenylmethyl radical and a Co(bpy)3]2+ cation, Crystal Engineering Communications, 6 (2004) 573 - 578 10. Maspoch D., et al., Carboxylic-substituted Polychlorotriphenylmethyl Radicals, New Organic Building-Blocks to Design Nanoporous Magnetic Molecular Materials, Comptes Rendus Chemie 8 (2005) 1213-1225 / This work shows the study of magnetism of different metal-organic materials from different families. On one side, there are the high spin molecules with negative uniaxial magnetic anisotropy. This anisotropy is the origin of an energy barrier in this type of molecules that blocks the orientation of magnetic moment in a single direction of the space. The magnetic moment can change the sense of its orientation in two ways: classically, by thermal activation over the energy barrier, or in a quantic manner, by quantic tunneling effect of the macroscopic magnetic moment through the energy barrier. This property allows thinking about applications of these type of molecules for ultra-high density magnetic recording and quantum computing, as magnetic qubits. The first part of this thesis is devoted to the study of the magnetic properties of some of this molecules of the Mn12 family, in different environments (from monocrystals to polymeric thin films and dissolutions), as well as new characterization techniques for the magnetic characterization of these molecules such as magnetic circular dichroism. This technique, allowed for the first time the measurement of some fundamental physics phenomena in this molecules through magneto-optical characterization, such as the quantum tunneling of the macroscopic magnetic moment. The second part of this thesis is devoted to the study of the magnetic properties of a new family of nanoporous metal-organic and pure organic materials, synthesized on the basis of perclorated triphenylmethyl radicals (PTM), functionalized with different carboxylic groups. The use of this radicals lead to nanoporous materials with a very high degree of porosity combined with interesting magnetic properties. For example, it is shown the first pure organic nanoporous ferromagnet, with a transition temperature of the order of 125 mK. The combination of these radicals with transition metal ions in the metal-organic approximation, also lead to interesting systems. The combination with Cu(III) ions with PTM radicals lead to the first nanoporous magnetic sponge, with reversible properties of solvent absorption-desorption that can be monitorized through its magnetic properties, thus becoming a low temperature magnetic sensor with solvent selectivity. Finally, the combination of Co(II) ions with PTM radicals lead to the firms nanoporous magnetic structure with non-interpenetrated helical networks that show an unusual slow relaxation of the magnetization; this relaxation shows a strong dependence with the magnetic field and can be described with the Davidson-Cole model.

Proteins are the most abundant biomolecules within a cell and are involved in all biochemical cellular processes ultimately determining cellular function. Therefore, to develop a complete understanding of cellular processes, obtaining knowledge about protein function and interaction at a molecular level is critical. Consequently, the investigation of proteins in their native environment or in partially purified mixtures is a major endeavour in modern life sciences. Due to their high chemical similarity, the inherent problem of studying proteins in complex mixtures is to specifically differentiate one protein of interest from the bulk of other proteins. Site-specific protein functionalisation strategies have become an indispensable tool in biochemical- and cell biology studies. This thesis presents the development of a new enzymatic site-specific protein functionalisation strategy that is based on the reversible covalent phosphocholination of short amino acid sequences in intact proteins. A synthetic strategy has been established that allows access to functionalised CDP-choline derivatives carrying fluorescent reporter groups, affinity tags or bioorthogonal handles. These CDP-choline derivatives serve as co-substrates for the bacterial phosphocholinating enzyme AnkX from Legionella pneumophila, which transfers a phosphocholine moiety to the switch II region of its native target protein Rab1b during infection. We identified the octapeptide sequence TITSSYYR as the minimum recognition sequence required to direct the AnkX catalysed phosphocholination and demonstrated the functionalisation of proteins of interest carrying this recognition tag at the N- or C-terminus as well as in internal loop regions. Moreover, this covalent modification can be hydrolytically reversed by the action of the Legionella enzyme Lem3, which makes the labeling strategy the first example of a covalent and reversible approach that is fully orthogonal to current existing methodologies. Thus, the here presented protein functionalisation approach holds the potential to increase the scope of possible labeling strategies in complex biological systems. In addition to the labeling of tagged target proteins, a CDP-choline derivative equipped with a biotin affinity-tag was synthesised and used in pull-down experiments to investigate the substrate scope of AnkX and to elucidate the role of protein phosphocholination during Legionella pneumophila infection. / Proteiner utgör huvudbeståndsdelen av alla biomolekyler i en cell. Dessa är involverade i alla cellulära processer som bestämmer cellens egenskaper. För att förstå de cellulära processerna är det nödvändigt att förstå proteinernas funktion på molekylär nivå. Att studera proteiner i deras naturliga omgivning, det vill säga inuti en cell eller i ett cellextrakt, är en stor utmaning i dagens livsvetenskaper. Eftersom proteiner är kemiskt lika varandra så är det svårt att skilja ett från tusentals andra. Att specifikt märka proteiner för att skilja ut dem från bakgrunden har blivit ett viktigt arbetssätt i modern biokemi och cellbiologi. Avhandlingen beskriver utvecklandet av en ny metod för reversibel och kovalent enzymatisk märkning baserat på fosfokolinering/defosfokolinering av en kort aminosyrasekvens i intakta proteiner. En syntesmetod för att framställa onaturliga CDP-kolinderivat har etablerats vilket tillåter oss att framställa CDP-kolin som bär en funktionalitet, vilket kan vara ett färgämne eller en affinitetstagg. Dessa onaturliga CDP-kolinderivat accepteras som co-substrat av enzymet AnkX från Legionella pneumophila vilket transfererar den funktionaliserade delen av CDP-kolinderivatet till en kort aminosyrasekvens baserad på AnkX’s naturliga substrat vid infektion, det lilla GTPaset Rab1. Under avhandlingsarbetets gång identifierades den kortaste aminosyrasekvensen som känns igen av AnkX, endast de åtta aminosyrorna TITSSYYR är nödvändiga för igenkänning av AnkX. Dessa åtta aminosyror kan genetiskt infogas i början, slutet eller mitt i ett protein för igenkänning och funktionalisering via AnkX och våra syntetiska CDP-kolinderivat. Vid Legionellainfektion i eukaryota celler klyvs fosfokolineringen efter en viss tid, eftersom Legionella pneumophila producerar ett fosfodiesteras, Lem3, som tar bort de fosfokolineringar som AnkX har installerat när de inte längre behövs. Vi har använt Lem3 för att ta bort märkning i sekvensen TITSS(PC)YYR, vilket gör vår strategi helt reversibel. Vi har kunnat demonstrera att AnkX-Lem3 systemet accepterar ett brett spektrum av CDP-kolinderivat, vilket gör metoden till den första av sitt slag, eftersom den är fullt reversibel. Vi har vidare undersökt vilka proteiner AnkX reagerar med inuti celler, vi använde oss av ett CDP-kolinderivat funktionaliserat med biotin, vilket har tillåtit oss att fiska ut alla de proteiner som fosfokolineras av AnkX. Förutom de små GTPaserna i Rab-familjen så identifierade vi även IMPDH2, ett enzym som reglerar det hastighetsbestämmande steget i syntesen av guanosin-nukleotider. Detta är mycket intressant, eftersom det leder till frågan ifall Legionella pneumophila manipulerar sin värdcell genom att förändra mängden GTP i förhållande till ATP.

Posttranslational modification (PTM) of proteins is essential to maintain homeostasis and viability in all eukaryotic cells. Hence, besides the sequence and 3D folding of a polypeptide, modification by multiple types of PTMs, ranging from small molecular groups to entire protein modules, adds another layer of complexity to protein function and regulation. The ubiquitin-like modifiers (UBLs) are such a group of evolutionary conserved protein modifiers, which by covalently conjugating to target proteins can modulate the subcellular localization and activity of their targets. One example of such a UBL, is the Ubiquitin related modifier 1 (Urm1). Since its discovery in 2000, Urm1 has been depicted as a dual function protein, which besides acting as a PTM, in addition functions as a sulfur carrier during the thio-modification of a specific group of tRNAs. Due to this dual capacity, Urm1 is considered as the evolutionary ancestor of the entire UBL family. At present, it is well established that Urm1, with help of its dedicated E1 enzyme Uba4/MOCS3, conjugates to multiple target proteins (urmylation) and that Urm1 thus plays important roles in viability and the response against oxidative stress. The aim of this thesis has been to, for the first time, investigate the role of Urm1 and Uba4 in a multicellular organism, utilising a multidisciplinary approach that integrates Drosophila genetics with classical biochemical assays and proteomics. In Paper I, we first characterized the Drosophila orthologues of Urm1 (CG33276) and Uba4 (CG13090), verified that they interact physically as well as genetically, and that they together can induce urmylation in the fly. By subsequently generating an Urm1 null Drosophila mutant (Urm1n123), we established that Urm1 is essential for viability and that flies lacking Urm1 are resistant to oxidative stress. Providing a molecular explanation for this phenotype, we demonstrated an involvement of Urm1 in the regulation of JNK signaling, including the transcription of the cytoprotective genes Jafrac1 and gstD1. Besides the resistance to oxidative stress, we have moreover (Manuscript IV) made an in-depth investigation of another phenotype displayed by Urm1n123 mutants, an overgrowth of third instar larval neuromuscular junctions (NMJs), a phenotype which is shared also with mutants lacking Uba4 (Uba4n29). To increase the understanding of Urm1 in the fly, we next employed a proteomics-based approach to identify candidate Urm1 target proteins (Paper II). Using this strategy, we identified 79 Urm1-interacting proteins during three different stages of fly development. Of these, six was biochemically confirmed to interact covalently with Urm1, whereas one was found to be associated with Urm1 by non-covalent means. In Manuscript III, we additionally identified the virally encoded oncogene Tax as a target of Urm1, both in Drosophila tissues and mammalian cell lines. In this study, we established a strong correlation between Tax urmylation and subcellular localization, and that Urm1 promoted a cytoplasmic accumulation and enhanced signalling activity of Tax, with implications for a potential role of Urm1 in Tax-induced oncogenesis. Taken together, this thesis provides a basic understanding of the potential roles and targets of Urm1 in a multicellular organism. The four studies included cover different aspects of Urm1 function and clearly points towards a highly dynamic role of protein urmylation in fly development, as well as in adult life.

Post-translational modifications (PTMs) of proteins can be a sign and/or cause of disease. These modified proteins have the potential to be used as biomarkers for diagnostic purposes. However, research in the field is limited. The challenge of having an accessible way of diagnosing patients in time and at a low cost is crucial to improve public health. Blood samples or other non-invasive methods to detect diseases such as Alzheimer’s disease, Parkinson’s disease, Amyotrophic lateral sclerosis and cancers are of urgent need. This report investigates PTMs as possible biomarkers measurable in biofluids, such as blood, for diagnosis and prognosis. Biomarkers like phospho-tau and amyloid-beta are examined in the context of neurodegenerative diseases, as well as phosphorylations on neurofilaments, TAR DNA-binding protein 43 and α-synuclein. All of these are detectable in blood. Several PTMs with connection to different types of cancers are also investigated, such as F3-phosphopeptide and AFP-L3. It was found that many biomarkers for the detection of cancers can potentially be found in extracellular vesicles in blood. Methods such as ELISA, PEA, SomaScan, xMAP, SIMOA and mass spectrometry (MS) are all now available on the market to quantify these PTMs. MS has revolutionized the fields of protein detection in the past and has further evolved to being capable of protein quantification. ELISA has been prevalent for decades and laid the groundwork for improved methods such as xMAP and SIMOA that are easy to use and provide adequate sensitivity. SomaScan and PEA lead the way in dynamic range and multiplexing capacity with around 7000 and 3000 protein assays. The soon-to-be-released technology NULISA, with promising values in sensitivity and dynamic range, is also investigated here. Additionally, a written ethical analysis regarding the process and consequences of biomarker quantification through these technologies was performed. Although the investigated biomarkers are detectable in biofluids, using them as clinical diagnostic markers still poses a challenge, which is why further research in the field is needed. Through an increased knowledge of PTMs of proteins and the right use of platforms, clinical diagnostics and population screenings can be done more efficiently improving public health around the world.

L'étude des modifications post-traductionnelles a connu au cours des dernières années un regain d'intérêt notable. Tout d'abord car elle s'effectue aujourd'hui au travers d'approches basées sur la spectrométrie de masse, technique qui pendant cette période a connu de profonds bouleversements, conduisant à des études plus aisées et systématiques.Mais aussi car tant par leur variété que par le rôle qu'elles jouent dans la vie et la régulation cellulaire, ces modifications ne peuvent plus être négligées. Par ailleurs au cours de ces quinze dernières années, nous avons assisté concernant les procaryotes à un changement total de paradigme. En effet à la fin des années 90, l'idée dominante était que ces modifications pouvaient exister chez ceux-ci mais de façon très partielle et/ou très particulière.Dans ce travail, les divers degrés d'iodation de la tyrosine ont été sondés par une approche de type «shotgun » sur un organe entier, la thyroïde de souris. L'efficacité de ce type d'approche démontrée, les modifications post-traductionnelles potentiellement présentes dans des organismes modèles radiorésistants, la bactérie Deinococcus deserti et l'archée Thermococcus gammatolerans ont été analysées. Dans le premier cas, les données de protéomique montrent que de nombreuses acétylations N-terminales portent sur un motif spécifique (essentiellement des thréonines et sérines), cas très atypique pour une bactérie. Chez Thermococcus gammatolerans les acétylations N-terminales sont rares, mais la présence d'acétylations sur les chaînes latérales des lysines est notable. La présence de phosphorylations sur ces mêmes protéines, laisse entrevoir un possible phénomène de « cross talk » entre les lysines acétylées et les sérines et/ou thréonines phosphorylées.Ici, nous démontrons que la complexité du protéome chez les procaryotes par le biais des MPT est bien réelle et que de possibles interdépendances entre MPT mériteraient un regard nouveau. / Recently, the study of post-translational modifications has greatly evolved, mainly because of crucial progresses in mass spectrometry methodology which have allowed high-throughput, high resolution analysis. Their variety and their role in the regulation of key molecular mechanisms are increasingly documented. In this work, the different degrees of iodination of tyrosine were probed with a "shotgun" approach carried out from an entire organ, the mice thyroid. Post-translational modifications present in two radioresistant organism models, the bacterium Deinococcus deserti and the archaeon Thermococcus gammatolerans, were analyzed. The large scale exploration of N-terminal acetylation in D. deserti indicates a specific pattern of this modification on serine and threonine, as well as an atypical, high propension to acetylation with 50% of modified N-termini. In T. gammatolerans, N-terminal acetylation is rare, but the presence of acetylation on lysine side chains is significant. The presence of phosphorylation on these proteins suggests a potential "cross talk" between the acetylated lysine and phosphorylated serine or threonine residues. This work demonstrates that the complexity of the proteome in prokaryotes through post-translational modifications is higher than expected when extremophiles are scrutinized compared to classical prokaryote models. Interdependencies between post-translational modifications definitively deserve a fresher look.

This paper deals with testing the method Reflectance Transformation Imaging (RTI) on the copy of the Piraeus-lion in Historic Museum in Stockholm. The purpose is to find out if it is possible to gather more information about the runic inscriptions. RTI is a method that records the surface normal of individual pixels in a digital photograph by analyzing the impact of light coming from different angles of entrance. RTI produces sort of a 3D-image of the object. There will be an overview of earlier interpretations of the runic scripts written on the lion as well as an overview of Varangians in the southeast. After examination of the lion with RTI, has a conclusion been made that the method should be performed again on similar items, and on the copy of the Piraeus-lion to evaluate its full potential.

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