<p>Endozepine is an alternative name for the testis-specific isoform of the acyl-CoA binding protein (t-ACBP). Acyl-CoA binding proteins form a highly conserved family of proteins, which bind long chain fatty acid esters with nanomolar affinity. They are also known to be endogenous ligands to the --amino butyric acid (GABA) receptor in the central nervous system and to play a role in a wide variety of cellular functions such as vesicular trafficking, fatty acid biosynthesis and gene regulation. A role for endozepine in apoptosis was suggested through promoter gene trapping studies using CHO22 cells in which 90 % reduction in the expression of endozepine correlated with delayed mitochondrial permeabilization, a reduced activation of caspase-3 (an activator of apoptosis) and a consequent resistance to C2-ceramide induced apoptosis.</p> <p>Transduction studies using Tat-GFP-ELP fusion protein showed that endozepine restored the sensitivity of mutant CHO22 cells to C2-ceramide induced apoptosis. In this thesis, we have investigated two hypotheses for the involvement of endozepine in ceramide-induced apoptosis. The first hypothesis is that endozepine contributes to apoptosis through the transport of palmitoyl-CoA, a substrate required for the de novo synthesis of ceramide. The second hypothesis is that endozepine interacts directly with ceramide leading to interaction with peripheral benzodiazepine receptor and a subsequent opening of the mitochondria permeability transition pore, leading to apoptosis.</p>

Un dels reptes més ambiciosos que té avui en dia la comunitat científica és millorar el coneixement de la reacció d’oxidació de l’aigua que dóna lloc a oxigen, protons i electrons. D’altra banda els epòxids son intermedis de reaccions diverses en la indústria química, particularment per a la síntesi de diversos polímers i “fine chemicals” com ara productes farmacèutics, additius alimentaris o fragàncies. Els aquo complexes mononuclears i dinuclears de ruteni amb lligands polipiridílics han estat estudiats a fons pel nostre grup de recerca i d’altres, convertint-se en catalitzadors útils per a l’oxidació d’aigua i per a l’epoxidació d’alquens. Aquesta tesi vol presentar i discutir els següents temes relacionat: 1. La immobilització de dos nous complexes moleculars del tipus “Ru-Hbpp” capaços d’oxidar aigua, sobre òxids metàl·lics com són el TiO2 i el SiO2 així com fent ús del polímer Nafion per tal d’assolir l’objectiu final de construir una cel·la foto-electroquímica. El comportament catalític d’aquests nous sistemes serà discutit i comparat tant amb les espècies homòlogues moleculars com amb d’altres sistemes similars. 2. La síntesi i caracterització d’un complex dinuclear de ruteni que conté un lligand pont amb disposició bis-facial. La seva reactivitat envers l’oxidació d’aigua i d’olefines i la discussió comparant amb sistemes ja descrits a la literatura també formen part d’aquesta secció. 3. En aquest capítol s’ha plantejat la síntesi d’una nova família de lligands híbrids tetradentats capaços de coordinar el ruteni tant per N com per C, camp en el que se n’han fet diversos avanços. Així mateix descrivim un nou complex dinuclear de ruteni contenint un d’aquests nous lligands. 4. La preparació i caracterització tant espectroscòpica com electroquímica d’una família de complexos que tenen com a fórmula general [RuII(PY4Im)(X)]n+ (X = Cl, n = 1 or X = H2O, n = 2), on PY4Im és el lligand pentadentat 1,3-bis(bis(2-piridil)metil)imidazol-2-ilidè. En aquest apartat també es descriurà l’estudi catalític d’aquestes noves espècies envers l’oxidació d’aigua i alquens. / The understanding of the water oxidation reaction to molecular oxygen is still one of the great challenges faced by scientific community. On the other hand the epoxidation of olefins is a reaction of high relevance in both industry and academia. Epoxides are important intermediates in the chemical industry, particularly for the synthesis of various polymers and fine chemicals, such as pharmaceuticals, food additives, or flavor and fragrance compounds. Mononuclear and dinuclear Ru aqua complexes containing polypyridylic ligans have been recently studied by ourselves and by other groups, and have become a very useful platform for both water oxidation catalysis and the epoxidation of alkenes. In this thesis we present and discuss the following related topics: 1. Immobilization of two new “Ru-Hbpp” molecular water oxidation catalysts onto metal oxides such as TiO2, SiO2 as well as into Nafion in order to succed in the building up of a photo-electrochemical cell. The catalytic performance of these new water oxidation materials will be discussed and compared with their homogeneous counterparts and other related systems previously reported. 2. Synthesis and characterization of a novel diruthenium complex containing the bis-facial hexadentate bridging ligandg Hbimp. Its reactivity towards the oxidation of water and olefins and the comparative discussion with the already reported family of related dinuclear complexes. 3. The synthesis of a new family of tetradentate bridging phthalazine-triazole ligands capable to coordinate a metal centre via both N and C atoms is planned and attempted. The synthesis and characterization of a new dinuclear Ru complex containing one of these ligands is here reported and further discussed. 4. The synthesis and the spectroscopical and electrochemical characterization of a new family of complexes with general formula [RuII(PY4Im)(X)]n+ (X = Cl, n = 1 or X = H2O, n = 2), where PY4Im is the pentadentate 1,3-bis(bis(2-pyridyl)methyl)imidazol-2-ylidene ligand. These results together with the performance of the new species towards the oxidation of water and alkenes will be thoroughly discussed.

The transition metal complexes cyclopentadienylrhenium tricarbonyl [CpRe(CO)3, Cp = cyclopentadienyl] and (isopropylcyclopentadienyl)rhenium tricarbonyl [(i-PrCp)Re(CO)3, i-Pr = isopropyl] were photolysed in alkanes at low temperature and the resulting alkane complexes, of the general formula Cp'Re(CO)2(alkane) (Cp' = Cp or (i-PrCp)), were studied using NMR spectroscopy. Characteristic proton chemical shifts (δ) and couplings (3JHH) were observed for alkane complexes of several linear, branched and cyclic alkanes of up to eight carbons. Alkanes with chemically distinct methyl (CH3) and/or methylene (CH2) units were observed alternatively binding through each unit to rhenium. No bound methine unit was observed. Large C-H coupling constants (1JCH) were observed for protons of several bound CH3 and CH2 units, indicating the bound C-H is intact. These species are, thus, alkane sigma (σ) complexes, wherein the alkane has an agostic (M-H-C, 3 centre 2 electron) interaction with the rhenium centre. The CH3 binding mode of (i-PrCp)Re(CO)2(1-pentane) was elucidated; sequential deuteration in the bound CH3 revealed an equilibrium isotope effect (EIE) in the remaining proton/s, confirming that only one C-H has an agostic interaction with rhenium at any instant . NMR parameters δ(1H) (-8.22), δ(13C) ( 42.4) and 1JCH (85 Hz) for the complexed C-H reveal it is unequivocally intact and yet strongly interacting with the rhenium centre, hallmarks for the agostic interaction. Intramolecular exchange was identified between pentane complex isomers Cp'Re(CO)2(1-pentane), Cp'Re(CO)2(2-pentane) and Cp'Re(CO)2(3-pentane). Equilibrium constants were determined, revealing a preference for CH2 binding over CH3. The inequivalent hydrogens found in methylene groups of cyclohexane at low temperature permitted simultaneous observation of axial and equatorial C-H protons of a bound CH2 in CpRe(CO)2(cyclohexane); an EIE, upon deuteration, indicated rapid exchange between complexed C-H bonds in the bound CH2 unit. The rhenium centre was found to prefer complexation of the axial C-H bond, over the equatorial, with K ~2.9. Intermolecular exchange of alkane ligands with free solvent was directly observed, in the competitive complexation of the [CpRe(CO)2] fragment to different alkanes in binary mixtures. The preference cyclohexane > cyclopentane > pentane > isobutane was established and equilibrium constants determined. The kinetics were followed by NMR and modelled, revealing rate constants; decay rates were also determined.

No description available.

Electron transport systems exist in the plasma membranes of all cells. Although not well characterised they play roles in cell growth and proliferation, hormone responses and other cell signalling events, but perhaps their most important role, especially in erythrocytes, is enabling the cell to respond to changes in both intra- and extracellular redox environments. Human erythrocytes possess a transmembrane electron transport capability that mediates the transfer of reducing equivalents from reduced intracellular species to oxidised extracellular species and is concomitant with proton extrusion. In the work for this thesis I showed that erythrocyte membranes contain a transmembrane WST-1 (water soluble tetrazolium-1) reductase activity that uses reducing equivalents from intracellular NADH to reduce extracellular WST-1. The rate of WST-1 reduction was increased by the presence of phenazine methosulfate and, although of low activity, it showed similar properties to a previously reported transmembrane NADH-oxidase activity. 1H NMR experiments showed that WST-1 was reversibly bound to the membrane and/or proteins in the membrane within the timeframe of the NMR experiment, confirming the location of the WST-1 reduction. Preliminary attempts to purify NADH:WST-1 reductase and NADH:ferricyanide reductase activities from the erythrocyte plasma membrane were inconclusive. The protein(s) responsible for the reduction of these oxidants appear to be of low abundance in the plasma membrane and may be part of a larger protein complex. Further work on the isolation of these redox activities is required before the protein(s) involved can be identified with any confidence. The ability of cells to export electrons suggests that an electron import mechanism might also exist to re-establish the cell�s redox-buffering equilibrium under conditions of oxidative stress. This hypothesis was tested in glucose-deprived erythrocytes using reduced glutathione and NADH as extracellular electron donors. It was shown that neither reduced glutathione nor NADH donated reducing equivalents through a transmembrane redox system. Extracellular NADH was, however, able to produce profound changes in starvation metabolism and methaemoglobin reduction rates. The addition of extracellular NADH caused a six-fold increase in the rate of lactate production above that observed in glucose-starved controls, together with a concomitant decrease in pyruvate production. In erythrocytes containing high levels of methaemoglobin, extracellular NADH increased the rate of methaemoglobin reduction in both the presence and absence of glucose. These results were explained by the leakage of lactate dehydrogenase from erythrocytes due to an admittedly low level of haemolysis. This caused the displacement of the intracellular pseudo-equilibrium of the lactate dehydrogenase reaction via transmembrane exchange of lactate, allowing the conversion of extracellular pyruvate to lactate and resulted in an increase in intracellular NADH concentrations. The latter increased the rate of methaemoglobin reduction. In conclusion, the work described in this thesis showed that erythrocyte membranes do not contain mechanisms for importing electrons or reducing equivalents from extracellular reduced glutathione or NADH. Erythrocytes do, however, contain an electron export system which can reduce extracellular oxidants such as WST-1 and the activity of this system depends on an intricate balance between intracellular antioxidants and enzyme activities. There is much still to be learnt about plasma membrane redox systems, little is known, for example, about the protein composition, mechanism of action, and the in vivo conditions under which these systems are most active.

Zugl.: Hannover, Univ., Diss., 2005

Zugl.: Berlin, Freie University, Diss., 2005.

Techn. University, Diss., 2006--München.

Würzburg, Universiẗat, Diss., 2007.

Techn. Hochsch., Diss.--Aachen, 2007.