While the catalytically powerful [Cu,Zn] superoxide dismutase (SOD1) possesses great potential as a therapeutic, unfavorable properties in circulation limit its use in clinical medicine. The small, water soluble dimer is rapidly excreted by the kidney. Previous initiatives have been used to increase the mass of the enzyme (PEGylation, liposome encapsulation). This has resulted in highly heterogeneous mixtures of modified SOD1, which are difficult to characterize. Furthermore, these modified proteins have utilized foreign material that has shown to elicit an inflammatory response. We developed an improved strategy that creates a homogenous high molecular weight SOD1 based on combinations of the protein itself. This was accomplished through the addition of a site-specific, azide functionalized cross-linker to unmodified SOD1, followed by the conjugation of SOD dimers using CuAAC and a bis-alkyne linker to form a 64 kDa SOD tetramer. The final product, bis-SOD, presents the fully catalytic activity of the combined proteins.

While the catalytically powerful [Cu,Zn] superoxide dismutase (SOD1) possesses great potential as a therapeutic, unfavorable properties in circulation limit its use in clinical medicine. The small, water soluble dimer is rapidly excreted by the kidney. Previous initiatives have been used to increase the mass of the enzyme (PEGylation, liposome encapsulation). This has resulted in highly heterogeneous mixtures of modified SOD1, which are difficult to characterize. Furthermore, these modified proteins have utilized foreign material that has shown to elicit an inflammatory response. We developed an improved strategy that creates a homogenous high molecular weight SOD1 based on combinations of the protein itself. This was accomplished through the addition of a site-specific, azide functionalized cross-linker to unmodified SOD1, followed by the conjugation of SOD dimers using CuAAC and a bis-alkyne linker to form a 64 kDa SOD tetramer. The final product, bis-SOD, presents the fully catalytic activity of the combined proteins.

The activity of superoxide dismutase (SOD) was determined in three kiwifruit (Actinidia) species including A. deliciosa, A. chinensis, and A. arguta. Among the species tested, the highest SOD activity was found in crude extracts prepared from fruit tissues of A. deliciosa. The highest enzyme activity was localized in seed, followed by locules, core and outer pericarp (OP). SOD activity in crude extract of whole fruit remained stable for at least one month when stored at -20℃. The effect of synthetic protease inhibitors (PI) on SOD activity was investigated. Supplementing crude kiwifruit extracts with PI improved SOD activity in freshly prepared extracts, and in extracts stored at 4℃, but had no effect on those stored at -20℃. Among the PI used, iodoacetamide (an inhibitor of cysteine proteases, for example, actinidin which is a principal protease found in kiwifruit) and PMSF (an inhibitor of serine proteases), had the most and least influence on SOD activity in crude kiwifruit extracts, respectively. There was a significant increase in SOD activity in kiwifruit (that were relatively firm) when the fruits were stored at low temperature (4℃). An increase in SOD activity was also correlated with a decrease in fruit firmness. Staining fruit tissues with nitroblue tetrazolium (NBT) provided evidence for stress-induced superoxide generation in kiwifruit tissues. Taken together, the changes in SOD activity and the capacity for stress-inducible superoxide production in post-harvest kiwifruit suggest that SOD might play a fundamental role in the storage life/ripening of kiwifruit.

Thesis (Ph. D.)--Virginia Polytechnic Institute and State University, 1992. / Vita. Abstract. Includes bibliographical references (leaf 193). Also available via the Internet.

Thesis (M. Sc.)--University of Canterbury, 2006. / Typescript (photocopy). Includes bibliographical references (leaves 86-97). Also available via the World Wide Web.

The production of reactive oxygen species by mitochondria is implicated in the mitochondrial dysfunction associated with a range of diseases and ageing. In contrast, reactive oxygen species produced by mitochondria are involved in redox signalling pathways necessary for modulating a number of cell processes. Mitochondrially targeted antioxidants comprised of an antioxidant moiety linked to a lipophilic triphenylphosphonium cation have recently been used to decrease reactive oxygen species-mediated oxidative damage to mitochondria and to investigate the role of mitochondrial reactive oxygen species in redox signalling. These lipophilic cations are selectively accumulated by mitochondria within cells due to the mitochondrial membrane potential. This thesis presents the synthesis and characterization of mitochondrially targeted antioxidant superoxide dismutase and thiol peroxidase mimetics. A mitochondrially targeted derivative of the Mn(II) macrocycle SOD mimetic M40403 (MitoSOD) was synthesized by Mn(II) template synthesis of a chiral tetraamine component and a triphenylphosphonium derivative of 2,6-pyridinedialdehyde. Racemic tetraamine was synthesized by mono-protection of racemic diamine followed by reductive amination of glyoxal and deprotection of di-protected tetraamine but overall this was found to be less efficient than a reported method based on trityl protection. The synthesis of the triphenylphosphonium derivative of 2,6-pyridinedialdehyde involved substitution of protected 4-bromo-2,6-pyridinedialdehyde by the thiolate of 3-mercaptoproanol followed by simultaneous deprotection and alkyl bromide formation, and triphenylphosphine substitution of the thioalkyl bromide substituent. MitoSOD was found to be more lipophilic than M40403 and was kinetically stable to dissociation to Mn(II) and macrocyclic ligand at physiological pH. Pulse radiolysis kinetic studies indicated both MitoSOD and M40403 catalyse the dismutation of superoxide. Fast conductivity and spectrophotometric measurements indicated the mechanism of catalysis involved reaction of the Mn(II) centre with superoxide to give a Mn(III)-peroxide intermediate which reacted with further superoxide to give the parent Mn(II) macrocycle. MitoSOD was significantly accumulated by mitochondria and this was dependent to some extent on the mitochondrial membrane potential. In addition, MitoSOD appeared to react with a product of mitochondrial succinate respiration. A mitochondrially targeted derivative of the organoselenium thiol peroxidase mimetic ebselen (Mitoebselen) was synthesized by O-alkylation of a phenolic ebselen derivative with a triphenylphosphonium derivative of an alkyl iodide. Reaction of excess triphenylphosphine with an ebselen derivative containing an alkyl iodide substituent resulted in substitution of iodide and, unexpectedly, reduction of the isoselenazole moiety to the diselenide redox form. Mitoebselen and its diselenide were both readily reduced to a selenol by an excess of the physiological thiol glutathione. Reaction of the selenol with excess peroxide generated the diselenide, possibly via reaction of unreacted selenol with Mitoebselen formed from a selenenic acid intermediate or with selenenic acid directly. Mitoebselen and its diselenide were both oxidized by excess peroxide to a selenoxide but these reactions were much slower than those between selenol and peroxides, and those between Mitoebselen or its diselenide with glutathione. Together these studies suggested cyclic pathways other than a selenolisoselenazole-selenol cycle could be involved in Mitoebselen or ebselen-catalysed thiol peroxidation.

No description available.

Thesis (M. Phil.)--University of Hong Kong, 2007. / Title proper from title frame. Also available in printed format.

Thesis (M.S. in zoology)--Washington State University, May 2010. / Title from PDF title page (viewed on June 22, 2010). "School of Biological Sciences." Includes bibliographical references (p. 35-39).

Thesis (M. Sc.)--University of Canterbury, 2006. / Typescript (photocopy). Includes bibliographical references (leaves 92-101). Also available via the World Wide Web.