<p>In today’s society there is an increasing need for energy, an increase which for the most part is supplied by the use of fossil fuels. Fossil fuel resources are limited and their use has harmful effects on the environment, therefore the development of technologies that produce clean energy sources is very appealing. Natural photosynthesis is capable of converting solar energy into chemical energy through a series of efficient energy and electron transfer reactions with water as the only electron source. Thus, constructing an artificial system that uses the same principles to convert sunlight into electricity or storable fuels like hydrogen is one of the major forces driving artificial photosynthesis research.</p><p>This thesis describes supramolecular complexes with the intention of mimicking the electron transfer reactions of the donor side in Photosystem II, where a manganese cluster together with a tyrosine catalyses the oxidation of water. All complexes are based on Ru(II)-trisbipyridine as a photosensitizer that is covalently linked to electron donors like tyrosine or manganese. Photochemical reactions are studied with time-resolved transient absorption and emission measurements. Electrochemical techniques are used to study the electrochemical behavior, and different photoelectrochemical techniques are used to investigate the complexes adsorbed onto titanium dioxide surfaces. In all complexes, intramolecular electron transfer occurs from the linked donor to photo-oxidized Ru(III). It is also observed that coordinated Mn(II) quenches the excited state of Ru(II), a reaction that is found to be distance dependent. However, by modifying one of the complexes, its excited state properties can be tuned in a way that decreases the quenching and keeps the electron transfer properties. The obtained results are of significance for the development of multinuclear Ru-Mn complexes that are capable of multi-electron transfer.</p>

A new strategy was developed to switch the luminescence of semiconductor quantum dots with chemical stimulations. It is based on the photoinduced transfer of either energy from CdSe-ZnS core-shell quantum dots to [1,3]oxazine ligands or electrons from the organic to the inorganic components. Upon addition of base or acid, energy or electron transfer pathways respectively become operative, leading to changes in the luminescence of the nanoparticles. These changes are fully reversible and can be exploited to probe the pH of aqueous solutions from 3 up to 11 and this design can lead to the development of pH-sensitive luminescent probes for biomedical applications based on the semiconductor quantum dots. Secondly, an operating principle to transduce the supramolecular association of complementary receptor-substrate pairs into an enhancement in the luminescence of sensitive quantum dots was identified. This system is based on the electrostatic adsorption of cationic quenchers on the surface of anionic quantum dots. The adsorbed quenchers efficiently suppress the emission character of the associated nanoparticles on the basis of photoinduced electron transfer. In the presence of target receptors able to bind the quenchers and prevent electron transfer, however, the luminescence of the quantum dots is restored. Thus, complementary receptor-substrate pairs can be identified with luminescence measurements relying on this system and this protocol can be adapted to signal protein-ligand interactions. Thirdly, a photochromic spiropyran with dithiolane appendage to adsorb on the surface of cadmium sulfide system was designed. The properties of the resulting photochrome-nanoparticle assemblies vary significantly with the experimental conditions selected for the preparation of the inorganic component. Finally, photochromic materials based on the photoinduced transfer of electrons from CdSe-ZnS core-shell quantum dots to bipyridinium dications were developed.

Though mass spectrometry has earned a central role in the field of proteomics due to its versatility in a wide range of experiments, challenges and complications are still encountered when using mass spectrometry to characterize protein structures, post-translational modifications (PTMs), and abundances. In this dissertation, analytical methods utilizing mass spectrometry have been developed to address challenges associated with both qualitative and quantitative protein characterization. The effectiveness of using multiple pepsin-like proteases, both separately and in mixtures, combined with online proteolysis using a special triaxial probe has been demonstrated on an amyloid beta peptide related to the onset of Alzheimer’s disease. These findings have broad implications in protein structural characterization studies using hydrogen-deuterium exchange mass spectrometry. A wider range of proteases (Lys-C, Glu-C, and trypsin) and multiple fragmentation methods (collisionally activated dissociation, electron transfer dissociation, and decision tree) have been utilized in the discovery-based PTM characterization of extracellular cellulosome proteins of the bioenergy-relevent organism Clostridium thermocellum, resulting in the identification of 85 previously unknown modification sites in 28 cellulosome proteins. These modifications may contribute to the structure and/or function of the cellulosome protein complex. By using peptide internal standards and a triple quadrupole mass spectrometer operating in selected reaction monitoring mode, a method has been developed for the absolute quantitation of the Clostridium thermocellum cellulosome protein machine in samples ranging in complexity from purified cellulosome samples to whole cell lysates as an alternative to a previously-developed enzyme-linked immunosorbent assay method of cellulosome quantitation. The precision of the cellulosome mass concentration in technical replicates is better than 5% relative standard deviation for all samples, indicating high precision of cellulosome mass concentration for this method. Though methods and results presented in this dissertation have implications in the study of Alzheimer’s disease and bioenergy research, more broadly this dissertation focuses on development of methods to contend with some of the more complex challenges associated with protein characterization currently presented to the field of proteomics.

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Les réactions de transfert de charge entre molécules sont des processus fondamentaux dans de nombreux domaines scientifiques. En particulier, elles sont responsables de la conversion de la lumière en énergie chimique (photosynthèse) ou en énergie électrique dans les cellules solaires organiques (domaine du photovoltaïque). Ces dernières visent à produire de l’électricité à coût réduit afin de contribuer aux besoins en énergie renouvelable. Leur rendement reste toutefois faible (environ 5-6% contre 25% pour les cellules à base de silicium) et leur amélioration requiert l’optimisation de chaque étape du processus de transformation de l’énergie lumineuse en courant électrique : absorption de la lumière ; dissociation des excitations ; séparation des charges ; transport des charges. Les processus de dissociation d’excitations, de transport de charge, ainsi que de recombinaison de charges (diminuant le rendement de la cellule solaire) impliquent tous des réactions de transfert d’électron. Les transistors à effet de champ sont des composants fondamentaux en électronique, utilisés comme interrupteurs, déterminant si le courant passe ou non. Les transistors organiques se posent comme alternative moins couteuse aux transistors à base de silicium. Comme dans les cellules solaires, les processus de transfert de charge sont d’une importance primordiale, gouvernant ici la mobilité des charges au sein du dispositif. Rendre les dispositifs d’électroniques organiques concurrentiels par rapport aux dispositifs inorganiques nécessite de les rendre plus performants. Cette amélioration des performances passe, entre autres, par une optimisation des processus de transfert de charge. Dans ce contexte, cette thèse porte sur une étude théorique, basée sur des méthodes de chimie quantique et de mécanique moléculaire, de systèmes donneur-pont-accepteur, dans le but de mieux comprendre les processus de transfert de charge au sein de ces systèmes, en vue de leur utilisation en électronique organique. Ces systèmes présentent plusieurs avantages pour les cellules solaires : le fait de lier chimiquement le donneur et l’accepteur réduit la ségrégation de phase et la séparation des charges est facilitée lorsque les charges sont formées loin l’une de l’autre (grâce à la présence du pont) car l’attraction de Coulomb entre charges générées est réduite. Des molécules pontées peuvent également avoir d’autres applications, comme par exemple l’amélioration de la mobilité des charges dans un transistor organique à effet de champ en permettant un transport entre couches moléculaires. Nous nous sommes intéressés dans un premier temps à des structures modèles afin de mieux comprendre les mécanismes fondamentaux associés aux processus à transfert de charge. Nous avons ensuite appliqué cette méthode à trois types de systèmes réels, synthétisés et caractérisés expérimentalement. La première étude concerne le transport de charge au sein d’un copolymère conjugué donneur/accepteur, F8BT. La seconde concerne l’utilisation de systèmes pontés en vue d’une utilisation dans des transistors à effets de champs organiques. La dernière concerne l’injection de charges dans un matériau organique conjugué via une couche mono-moléculaire auto-assemblée. Pour terminer, nous nous sommes intéressés à l’influence de la dynamique structurale (vibrations moléculaires) sur les vitesses des réactions de transfert de charge dans des systèmes donneur-pont-accepteur.

Electron transfer phenomena in proteins represent one of the most common types of biochemical reactions. They play a central role in energy conversion pathways in living cells, and are crucial components in respiration and photosynthesis. These complex biochemical reaction cascades consist of a series of proteins and protein complexes that couple a charge transfer to different forms of chemical energy. The efficiency and sophisticated optimisation of signal transfer in these natural redox chains has inspired engineering of artificial architectures mimicking essential properties of their natural analogues. Implementation of direct electron transfer (DET) in protein assemblies was a breakthrough in bioelectronics, providing a simple and efficient way for coupling biological recognition events to a signal transducer. DET avoids the use of redox mediators, reducing potential interferences and side reactions, as well as being more compatible with in vivo conditions. However, only a few haem proteins, including the redox protein cytochrome c (cyt.c), and blue copper enzymes show efficient DET on different kinds of electrodes. Previous investigations with cyt.c have mainly focused on heterogeneous electron transfer of monolayers of this protein on gold. An important advance was the fabrication of cyt.c multilayers by electrostatic layer-by-layer self-assembly. The ease of fabrication, the stability, and the controllable permeability of polyelectrolyte multilayers have made them particularly attractive for electroanalytical applications. With cyt.c and sulfonated polyaniline it was for the first time possible that fully electro-active multilayers of the redox protein could be prepared. This approach was extended to design an analytical signal chain based on multilayers of cyt.c and xanthine oxidase (XOD). The system does not need an external mediator but relies on an in situ generation of a mediating radical and thus allows a signal transfer from hypoxanthine via the substrate converting enzyme and cyt.c to the electrode. Another kind of a signal chain is based on assembling proteins in complexes on electrodes in such a way that a direct protein-protein electron transfer becomes feasible. This design does not need a redox mediator in analogy to natural protein communication. For this purpose, cyt.c and the enzyme bilirubin oxidase (BOD, EC 1.3.3.5) are co-immobilized in a self-assembled polyelectrolyte multilayer on gold electrodes. Although these two proteins are not natural reaction partners, the protein architecture facilitates an electron transfer from the electrode via multiple protein layers to molecular oxygen resulting in a significant catalytic reduction current. Finally, we describe a novel strategy for multi-protein layer-by-layer self-assembly combining cyt.c with an enzyme sulfite oxidase (SOx) without use of any additional polymer. Electrostatic interactions between these two proteins with rather separated pI values during the assembly process from a low ionic strength buffer were found sufficient for the layer-by-layer deposition of the both biomolecules. It is anticipated that the concepts described in this work will stimulate further progress in multilayer design of even more complex biomimetic signal cascades taking advantage of direct communication between proteins. / Elektronentransferphänomene in Proteinen stellen den häufigsten Typ biochemischer Reaktionen dar. Sie spielen eine zentrale Rolle bei der Energieumwandlung in der Zelle und sind entscheidende Komponenten in der Atmung und Photosynthese. Diese komplexen Kaskaden biochemischer Reaktionen setzen sich aus einer Reihe von Proteinen und Proteinkomplexen zusammen, die den Energietransfer an verschiedene Formen chemischer Energie koppeln. Die große Effektivität und Selektivität des Signaltransfers in diesen natürlichen Redoxketten war Vorbild für die Entwicklung künstlicher Architekturen, die die wesentlichen Eigenschaften ihrer natürlichen Analoga nachahmen. Die Implementierung des direkten Elektronentransfers (DET) von Proteinen mit Elektroden war ein Durchbruch im Bereich der Bioelektronik. Sie lieferte einen einfachen und effizienten Weg für das Koppeln biologischer Erkennungsereignisse an einen Signalumwandler. Durch den DET können Redoxmediatoren vermieden und damit potentielle Grenzflächen und Nebenreaktionen reduziert werden. Ebenso wird damit die Kompatibilität für in vivo Bedingungen erhöht. Jedoch zeigen nur einige Hämproteine wie das Redoxprotein Cytochrom c (Cyt c) und blaue Kupferproteine einen effizienten DET auf verschiedenen Elektrodentypen. Bisherige Untersuchungen mit Cyt c konzentrierten sich hauptsächlich auf den heterogenen Elektronentransfer von Monoschichten dieses Proteins auf Gold. Ein wichtiger Fortschritt war die Herstellung von Cyt c Multischichten durch die elektrostatische Layer-by-Layer-Technik. Die einfache Herstellung, die Stabilität sowie die kontrollierbaren Permeationseigenschaften von Polyelektrolyt-Multischichten machte sie besonders attraktiv für elektroanalytische Anwendungen. So gelang es auch zum ersten Mal vollständig elektroaktive Multischichten aus Cyt c und Polyanilinsulfonsäure zu präparieren. Dieser Ansatz wurde hier erweitert, um eine analytische Signalkette auf der Basis von Multischichten aus Cyt c und Xanthinoxidase zu entwerfen. Das System bedarf keinen externen Mediator, es hängt jedoch von der in situ Generierung eines vermittelnden Radikals ab und erlaubt daher einen Signaltransfer von Hypoxanthin über ein substratumwandelndes Enzym und Cyt c zur Elektrode. Eine andere Art von Signalketten basiert auf der Assemblierung von Proteinen in Komplexen auf Elektroden in solcher Art und Weise, daß ein direkter Protein-Protein-Elektronentransfer möglich wird. Dieser Ansatz benötigt keinen Redoxmediator in Analogie zu Beispielen aus dem biologischen Signaltransfer. Zu diesem Zweck werden Cyt c und das Enzym Bilirubinoxidase mit einem selbst-assemblierenden Polyelektrolyten auf einer Goldelektrode koimmobilisiert. Obwohl diese zwei Proteine keine natürlichen Reaktionspartner sind, unterstützt die Protein-Architektur einen Elektronentransfer von der Elektrode über mehrere Proteinschichten zu molekularem Sauerstoff und ergibt einen signifikanten katalytischen Reduktionsstrom. Schließlich wird eine neue Strategie beschrieben für eine Selbstassemblierung von Proteinen ohne zusätzlichen Polyelektrolyten - am Beispiel der Kombination von Cyt c mit Sulfitoxidase. Es stellte sich heraus, daß die elektrostatische Wechselwirkung zwischen diesen zwei Proteinen mit ziemlich weit voneinander entfernt liegenden pI-Werten während des Assemblierungsprozesses durch einen Puffer mit geringer Ionenstärke ausreicht um die beiden Biomoleküle nach dem Layer-by-Layer-Prinzip auf einer Elektrode abzuscheiden. Es wird erwartet, daß das entwickelte Konzept von Multiprotein-Assemblaten auf Elektroden weitere Fortschritte bei dem Entwurf von Multischichten und sogar noch komplexeren biomimetischen Signalkaskaden anregen wird und dabei der Vorteil der direkten Kommunikation zwischen Proteinen genutzt wird.

First studies of electron transfer in [N]phenylenes were performed in bimolecular quenching reactions of angular [3]- and triangular [4]phenylene with various electron acceptors. The relation between the quenching rate constants kq and the free energy change of the electron transfer (ΔG0CS ) could be described by the Rehm-Weller equation. From the experimental results, a reorganization energy λ of 0.7 eV was derived. Intramolecular electron transfer reactions were studied in an [N]phenylene bichomophore and a corresponding reference compound. Fluorescence lifetime and quantum yield of the bichromophor display a characteristic dependence on the solvent polarity, whereas the corresponding values of the reference compound remain constant. From the results, a nearly isoenergonic ΔG0CS can be determined. As the triplet quantum yield is nearly independent of the polarity, charge recombination leads to the population of the triplet state.

Photosystem II is unique! It remains the only enzyme that can oxidize water using light as energy input. Water oxidation in photosystem II is catalyzed by the CaMn4 cluster. The electrons extracted from the CaMn4 cluster are transferred to P680+ via the tyrosine residue D1-Tyr161 (YZ). Favorable oxidation of YZ is coupled to a proton transfer along a hydrogen bond to the nearby D1-His190 residue, resulting in the neutral radical YZ•. By illuminating photosystem II at cryogenic temperatures, YZ• can be trapped in a stable state. Magnetic interaction between this radical and the CaMn4 cluster gives rise to a split electron paramagnetic resonance (EPR) signal with characteristics that depend on the oxidation state (S state) of the cluster. The mechanism by which the split EPR signals are formed is different depending on the S state. In the S0 and S1 states, split signal induction proceeds via a P680+-centered mechanism, whereas in the S2 and S3 states, our results show that split induction stems from a Mn-centered mechanism. This S state-dependent pattern of split EPR signal induction can be correlated to the charge of the CaMn4 cluster in the S state in question and has prompted us to propose a general model for the induction mechanism across the different S states. At the heart of this model is the stability or otherwise of the YZ•–(D1-His190)+ pair during cryogenic illumination. The model is closely related to the sequence of electron and proton transfers from the cluster during the S cycle. Furthermore, the important hydrogen bond between YZ and D1-His190 has been investigated by following the split EPR signal formation in the different S states as a function of pH. All split EPR signals investigated decrease in intensity with a pKa of ~4-5. This pKa can be correlated to a titration event that disrupts the essential hydrogen bond, possibly by a direct protonation of D1-His190.  This has important consequences for the function of the CaMn4 cluster as this critical YZ–D1-His190 hydrogen bond steers a multitude of reactions at the cluster.

In the society of today the need for alternative energy sources is increasing. The construction of artificial devices for the conversion of sunlight into electricity or fuel seems very attractive from an environmental point of view, since these devices are based on processes that does not necessarily generate any harmful biproducts. In the oxygen evolving photosynthetic process highly efficient energy and electron transfer reactions are responsible for the conversion of the sunlight into chemically stored energy and if the same principles can be used in an artificial device, the only electron supply required, is water. This thesis describes energy and electron transfer reactions in supramolecular complexes where the reactions are intended to mimic the basic steps in the photosynthetic process. All complexes are based on ruthenium(II)-trisbipyridine as photosensitizer, that is covalently linked to electron donors or electron or energy acceptors. The photochemical reactions were studied with time resolved transient absorption and emission measurements. In the complexes that mimic the donor side of Photosystem II, where a manganese cluster together with tyrosine catalyses the oxidation of water, intramolecular electron transfer was found to occur from Mn(II) or tyrosine to photo-oxidized Ru(III). Studies of a series of Ru(II)-Mn(II) complexes gave information of the quenching of the Ru(II) excited state by the coordinated Mn(II), which is important for the development of multi-nuclear Ru(II)-Mn complexes. In the supramolecular triad, PTZ-Ru2+-Q, the charge separated state, PTZ+●-Ru2+-Q-●, was rapidly formed, and further development where a second electron acceptor is linked to quinone is planned. Ultra fast energy transfer τ&lt;200 fs), was obtained between ruthenium(II) and osmium(II) in a small artificial antenna fragment. Fast and efficient energy transfer is important in larger antennas or photonic wires where a rapid energy transfer is desired over a large distance.

In today’s society there is an increasing need for energy, an increase which for the most part is supplied by the use of fossil fuels. Fossil fuel resources are limited and their use has harmful effects on the environment, therefore the development of technologies that produce clean energy sources is very appealing. Natural photosynthesis is capable of converting solar energy into chemical energy through a series of efficient energy and electron transfer reactions with water as the only electron source. Thus, constructing an artificial system that uses the same principles to convert sunlight into electricity or storable fuels like hydrogen is one of the major forces driving artificial photosynthesis research. This thesis describes supramolecular complexes with the intention of mimicking the electron transfer reactions of the donor side in Photosystem II, where a manganese cluster together with a tyrosine catalyses the oxidation of water. All complexes are based on Ru(II)-trisbipyridine as a photosensitizer that is covalently linked to electron donors like tyrosine or manganese. Photochemical reactions are studied with time-resolved transient absorption and emission measurements. Electrochemical techniques are used to study the electrochemical behavior, and different photoelectrochemical techniques are used to investigate the complexes adsorbed onto titanium dioxide surfaces. In all complexes, intramolecular electron transfer occurs from the linked donor to photo-oxidized Ru(III). It is also observed that coordinated Mn(II) quenches the excited state of Ru(II), a reaction that is found to be distance dependent. However, by modifying one of the complexes, its excited state properties can be tuned in a way that decreases the quenching and keeps the electron transfer properties. The obtained results are of significance for the development of multinuclear Ru-Mn complexes that are capable of multi-electron transfer.