This thesis focuses on amyloid proteins, a class of proteins that convert into amyloid fibrils. Such proteins are of high interest because they are related to many of the neurodegenerative diseases... Show moreThis thesis focuses on amyloid proteins, a class of proteins that convert into amyloid fibrils. Such proteins are of high interest because they are related to many of the neurodegenerative diseases. In the brains of patients with neurodegenerative diseases, plaques of β-sheet amyloid aggregates are found, but the mechanism of their formation and their role vis-à-vis the disease are unknown. Aggregation is difficult to study because amyloids are intrinsically disordered proteins that lack an ordered structure in solution. Here we apply electron paramagnetic resonance (EPR) as a new technique to better understand the properties of amyloid oligomers and their formation. Show less
Decentralized plug and play systems for energy production are the future picture of our society. Artificial photosynthetic systems are used for this purpose. These systems are inspired by natural... Show moreDecentralized plug and play systems for energy production are the future picture of our society. Artificial photosynthetic systems are used for this purpose. These systems are inspired by natural photosynthesis.Chapter 2 illustrates the detailed construction of a three-flash flow cell to study light induced biological reactions, specifically the water oxidation reaction of Photosystem II. This instrument is capable of producing the S2+, S3+ and S0n intermediate states of the Kok cycle. I confirmed these by EPR experiments.In chapter 3, I study the mechanism of the major LHC II involved in conformational switching from light harvesting to the photoprotective state, in which excess light is dissipated as heat. I use MAS NMR as a non-invasive method to understand the structure and environment around Arg residues in Chlamydomonas reinhardtii LHC II. In this approach the Arg amino acid is selectively labeled with 13C isotopesChapter 4 studies the structure of self-aggregated semi synthetic Zn 31 amino chlorin that forms syn-anti parallel stacks according to the solid state NMR data. The small change of removing 33 methyl group of Zn 31 aminomethyl chlorin has significant effect on self-assembly. Finally, chapter 5 provides an outlook on future research. Show less
Ever since the structural data of biological macromolecules became available, there has been consistent struggle to relate this new information to the existing spectroscopy, activity and... Show moreEver since the structural data of biological macromolecules became available, there has been consistent struggle to relate this new information to the existing spectroscopy, activity and theoretical descriptions of these proteins and to understand the evolution and/or to predict the role of yet uncharacterized gene products in this light. The research presented in this thesis primarily deals with understanding the structure__function relationship of a newly discovered blue copper protein. The protein is derived from Streptomyces coelicolor and is called small laccase (SLAC). It utilizes four copper ions to catalyze the oxidation of substrate molecules concomitant with the reduction of oxygen to water. The catalytic cycle of this enzyme is studied using a variety of spectroscopic and kinetic methods in an attempt to improve our understanding of the internal operations which are critical to its functioning. The new results obtained are presented in this thesis. Show less
Spin labeling and electron paramagnetic resonance (EPR) have been employed to study structure and dynamics of proteins. The surface polarity of four single cysteine mutants of the Zn-azurin in... Show moreSpin labeling and electron paramagnetic resonance (EPR) have been employed to study structure and dynamics of proteins. The surface polarity of four single cysteine mutants of the Zn-azurin in frozen solution were studied using 275 GHz EPR (J-band), with the advantage compared to 9 GHz (X-band) and 95 GHz (W-band) to allow differentiating between sites having small differences in polarity. The polarity/proticity profiles of the four mutants have been obtained and compared with data already reported in literature of spin labels in different solvents. In order to detect distances at the nm scale, azurin was used as a known model system for double electron-electron spin resonance. The conformation of the spin label linker was modelled. The model was found to be in good agreement with experimental results and can be extended to other proteins. The dynamics of the transient complex of Nostoc sp. PCC 7119 cytochrome f __ plastocyanin was investigated by NMR using the paramagnetic relaxation enhancements. The experimental distance restraints, used in docking calculations, are best interpreted by the presence of a dynamic ensemble of protein-protein orientations within the complex, rather than by a single, well-defined structure, and can be described with an encounter complex model. Show less
Verschillende biologische systemen, zoals het 50S ribosomale Hsp15 complex, een schistosomiasis diagnostisch antigen bindend antilichaam fragment (Fab 54-5C10-A), het zuurstof producerende enzym... Show moreVerschillende biologische systemen, zoals het 50S ribosomale Hsp15 complex, een schistosomiasis diagnostisch antigen bindend antilichaam fragment (Fab 54-5C10-A), het zuurstof producerende enzym chloriet dismutase (Cld) en het belangrijke regulatoreiwit van microtubule dynamiek (EB1) zijn onderzocht met geavanceerde biochemische technieken. De structurele informatie verkregen met electronenmicroscopie (EM), kleine hoek verstrooiing van R_ntgenstraling (SAXS) en R_ntgendiffractie aan individuele eiwitkristallen werd aangevuld met informatie uit andere biofysische en biochemische methoden, zoals kolom chromatografie, gel elektroforese, oppervlakte plasmon resonantie (SPR), massa spectrometrie (MS) en electron paramagnetische resonantie (EPR). Hoewel de in dit proefschrift beschreven biologische systemen uiteenlopen wat betreft chemische samenstelling, oorsprong en biologische functie, is er juist ook gekeken naar een gemeenschappelijk kenmerk: ge_nduceerde conformatieveranderingen binnen het eiwit, eiwit-RNA of eiwitcarbohydraat complex. Show less
A spectrometer for Electron Paramagnetic Resonance (EPR), operating at the very high microwave frequency of 275 GHz in both continuous wave (cw) and pulsed mode, is described and its capabilities... Show moreA spectrometer for Electron Paramagnetic Resonance (EPR), operating at the very high microwave frequency of 275 GHz in both continuous wave (cw) and pulsed mode, is described and its capabilities are illustrated. A superconducting magnet provides the magnetic field up to 12 Tesla and experiments are possible between room temperature and 5 K. The system utilizes pseudo-optical techniques to minimize microwave losses and a single-mode resonator is used to maximize the microwave field at the sample position. The material under study can also be subjected to a radiofrequency radiation field to enable Electron Nuclear Double Resonance (ENDOR) experiments. Technical details of the design and the operation are presented. First results are discussed which demonstrate the increased sensitivity and resolution compared to experiments at lower microwave frequencies. These concern various materials, ranging from semi-conductors to proteins, including an aqueous solution of a proxyl radical at room temperature. ENDOR performance is illustrated with data from a sample of zinc-germanium-phosphide (ZnGeP2) doped with Manganese (Mn). Separate chapters cover experiments on the organic semiconductor P3HT and the observation of Dynamic Nuclear Polarization (DNP) in zinc oxide (ZnO). Show less
Plants harvest light energy and convert it into chemical energy. Light absorption by photosystems I and II (PSI and PSII) results in charge separations in their reaction centers (RCs), initiating a... Show morePlants harvest light energy and convert it into chemical energy. Light absorption by photosystems I and II (PSI and PSII) results in charge separations in their reaction centers (RCs), initiating a chain of redox reactions with PSI generating the reducing power for CO2 assimilation into sugars, and PSII oxidizing the ultimate electron donor, H2O, to O2. The rates of PSI and II must be balanced. If PSII releases more electrons than PSI can use, the intermediary pool of plastoquinone molecules and the primary acceptor quinone QA become fully reduced. PSII charge separation then leads to recombination, whereby the RC triplet state and hence singlet oxygen can be formed, causing damage to PSII that can decrease the photosynthetic yield. Photoinhibition results. Charge recombination to the triplet state may probably be avoided to some extent by a cyclic electron transfer (CET) pathway, short-circuiting the charge separation, which is somehow switched on when needed by cytochrome b559, an intrinsic subunit of PSII. Mechanism, quantitative significance, and even the very existence of CET continue to be subject to debate, however. This thesis describes studies on triplet generation, cytochrome b559, CET induction conditions, and other aspects of this elusive protection mechanism against photoinhibition. Show less
Ubbink, M.; Warmerdam, G.C.M.; Campos, A.P.; Teixeira, M.; Canters, G.W. 1994
