Tyrosinase (E.C. 1.14.18.1) is a key enzyme in the biosynthesis of melanin. The control of melanin sythesis was explored in skin melanocytes of the following strains; wild type (C57BL/6J-C/C) (which maximally synthesize melanin at normal mammalian body temperature, Himalayan (C57BL/6J-cʰ/cʰ) (which maximally synthesize melanin at temperatures below 37°C) and albino (Balb c-c/c) (a mutant which does not synthesize melanin) The effect of a-MSH on tyrosinase activity was initially investigated. A skin culture tyrosinase assay that made it possible to measure the effect of α-MSH on the activity of this enzyme in vitro was first developed. It was found that α-MSH activated the wild type and Himalayan tyrosinase in a dose-dependent manner and that this activation did not require the de novo synthesis of new enzyme. The role of glycosylation on the wild type and particularly the Himalayan tyrosinase activity was next investigated. The results do not support, but are not in conflict with the theory that the Himalayan tyrosinase is inherently underglycosylated. Translation and transcription as additional control mechanisms of tyrosinase activity was finally investigated. The correlation between the levels of tyrosinase activity, abundance of the enzyme and the synthesis of tyrosinase mRNA in wild type, Himalayan and albino mice was determined. It was shown that the levels of newly synthesized tyrosinase and tyrosinase mRNA transcripts were higher in the wild type than in the Himalayan skin. This could account for the reduced tyrosinase activity in the Himalayan mutant at normal body temperature. Low levels of tyrosinase mRNA were found in the albino skin though there was no immunodetectable enzyme in this tissue.

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第21276号 / 工博第4504号 / 新制||工||1700(附属図書館) / 京都大学大学院工学研究科合成・生物化学専攻 / (主査)教授 跡見 晴幸, 教授 森 泰生, 教授 梅田 眞郷 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DGAM

No description available.

No description available.

No description available.

<p> DL-Mimosine has been synthesized by debenzylation and detosylation of the product obtained by condensation of 3-benzyloxy-4-pyrone with β-amino-α-tosylaminopropionic acid. A new method for the isolation of mimosine from Leucaena glauca Benth. is described. The biosynthesis of mimosine was studied by feeding radioactive aspartates, glycerol, glycerate and ribose to Mimosa pudica L. . Mimosine-C^14 was isolated and partially degraded, and it was shown that the carbon-3 of aspartic acid is specifically incorporated into the pyridone ring.</p> / Thesis / Doctor of Philosophy (PhD)

No description available.

No description available.

Diabetes mellitus dramatically increases the risk for atherosclerotic cardiovascular disease. It has been established that chronic hyperglycemia promotes an increase in glucose flux through the hexosamine biosynthesis pathway (HBP). Central to this pathway is glutamine:fructose-6-phosphate amidotransferase (GFAT), the rate-limiting enzyme controlling the conversion of glucose to glucosamine. We have shown that glucosamine is a potent inducer of endoplasmic reticulum (ER) stress, which is characterized by the accumulation of misfolded proteins in the ER. Chronic ER stress can initiate a multifaceted response that results in lipid accumulation, inflammation and apoptosis: the hallmark features of atherosclerosis. We hypothesized that conditions of chronic hyperglycemia, associated with diabetes mellitus, can accelerate the development of atherosclerosis by a mechanism that involves increased HBP flux resulting in glucosamine-induced ER stress and the subsequent activation of pro-atherogenic pathways. In support of the hypothesis we found that glucosamine-supplemented apoE-/- mice had elevated levels of ER stress and atherosclerosis. Mechanistically, our data showed that glucosamine induced ER stress by interfering with the lipid-linked oligosaccharide biosynthesis pathway and protein N-glycosylation. These findings support a model by which conditions of hyperglycemia promote vascular complications through a glucosamine-intermediate. / Thesis / Doctor of Philosophy (PhD) / Diabetes mellitus dramatically increases the risk for heart attacks and strokes. High blood glucose is utilized in cells through its conversion into metabolites, such as glucosamine. We hypothesized that conditions of high blood glucose can led to an increase in intracellular glucosamine which can initiate pathways involved in accelerating atherosclerosis. Our results show that this is possible in both human cells and mice.

<p> The increase in microbes resistant to a wide array of antibiotics has led to the need for the development of novel antimicrobials. However in order to develop new antimicrobials, novel pathways need to be targeted. Teichoic acid is an anionic polymer covalently attached to the cell wall of Gram-positive bacteria. Recent research has demonstrated that teichoic acid genes are indispensable to the viability of Bacillus subtilis. This makes teichoic acid biosynthetic proteins ideal candidates for the development of a new antimicrobial. Of the teichoic acid glycerol phosphate (tag) genes involved in the biosynthesis of teichoic acid in B. subtilis 168, a conditional deletion mutant of tagD, whose protein product encodes the proposed glycerol-3-phosphatecytidylyltransferase, has been previously constructed and was shown to have a lethal phenotype upon depletion of TagD. This was used in a microarray analysis to find genes that were transcriptionally up-regulated upon the depletion of TagD in B. subtilis 168. Ten candidate genes were selected from those up-regulated and used in the design of a novel, real-time, cell-based luminescent reporter system that responds to lesions in wall biosynthesis. Characterization of these reporter systems in tag gene deletion backgrounds and an examination of their response to antibiotics of various mechanism of action led to the identification of our candidate reporter system P ywac, a robust reporter of both lesions in teichoic acid and peptidoglycan synthesis. In a proof-of-principle screen, the use of Pywac as a reporter of lesions in the cell wall was validated. This reporter system is unique in that it combines conventional genetics with a high throughput capacity. It will not only be amenable for screening small molecules to find inhibitors that impinge on teichoic acid biosynthesis, but it can also be used to probe genetic interactions in B. subtilis. </p> / Thesis / Master of Science (MSc)