The addition of Mn(II) to an early stationary-phase Deinococcus radiodurans RI culture could induce a new round of cell division (MnCD effect). The addition of Mn(II) could also stimulate the utilization of glucose and fructose in this bacterium. Class II fructose-1,6-bisphosphate aldolase (FBA) is an Mn-dependent key enzyme in pentose phosphate pathway. Therefore, in this research, we focused on the studies of the fba gene. Base on the gene sequence, FBA protein was composed of 306 amino acids, (M.W., 32.4 kDa¡F pI, 5.4). The expected PCR product size of the fba gene is 9.3 kbp. We had amplified the fba gene by using both Taq DNA polymerase and pfu turbo DNA polymerase. The sequence of the pfu turbo DNA polymerase products showed a higher homology with the fba gene than those of using Taq DNA polymerase. These amplified fba gene was cloned into three expression vectors, pGEX-4T-2, pQE30, and pET28a, and then further expressed in E. coli BL21(DE3)RIL and JM109. The recombinant GST-FBA protein could be overproduced in pTDA2/BL21(DE3)RIL. However, the expressed insoluble protein accumulated as inclusion bodies in the cells and exhibited no enzyme activity. After partial purification, and processing by thrombin protease cleavage, urea treatment, and the addition of Mn(II), this enzyme still showed no activity. The recombinant pEDA2/BL21(DE3)RIL strain cells grew in 18¢J and induced by 0.1mM IPTG could produced a soluble form His-Thrombin-T7-FBA protein which performed a 50X higher activities than those cells grew in 30¢J. This result indicated that decreasing the indicatioin temperature could improve the protein solubility and activity.

Sanguinolaria rostrata is a deep site-burrowing bivalve commonly found in the southwest coastal waters of Taiwan. It has a long siphon extending to the surface. It is reported that exposed S. rostrata dies in few days without silt. However trussing it up rubber bands or cut off the hinge, it can survive for over one month in the laboratory. In this study the relation of shell closing and mortality for S. rostrata were examined and compared with the hard clam Meretrix lusoria in the similar environments. The size of adductor muscle and its ratio to the shell size, the strength of shell closing and the tissue structure of adductor muscle were examined. The quantities of fructose 2,6- biphosphate, an intermediate of glycolysis, in the adductor muscle of S. rostrata were determined. The results indicated that the average strength of closing shell for S. rostrata was 36.65% and for M. lusoria was 41.19%. The trends of tropic shell closing strength and the size of adductor muscle as well as shell closing strength and the adductor muscle wet weight were the same for the two species. The ranges of strength for muscle closing among S. rostrata of different sizes were smaller than those of M. lusoria. The average ratio of the adductor muscle microfiber to muscle was 55.6¢Mfor S. rostrata and 83.2¢M for M. lusoria. Therefore, the adductor muscle of S. rostrata is looser to M. lusoria. The concentration of fructose 2,6- biphosphate fluctuated widely to the unclamped S. rostrata in the first 6 hours and the concentration reached 7.58£gmole/mg at most. The concentration did not rise between 6 and 24 hours, indicating that unclamped S. rostrata consumed energy within the first 6 hours, then showed no sign of consuming energy.

The equilibria of adsorption on calcium and hydrogen forms of zeolite Y by equimolar solutions of 12.5 %, 20%, 25%, 30% and 35% w/v of mixtures of glucose, G and fructose, F / also the non-equimolar mixtures of 20% w/v glucose - 30% w/v fructose, 30% w/v glucose - 20% w/v fructose, 25% w/v glucose &ndash / 35% w/v fructose, and 35% w/v glucose-25% w/v fructose solutions, which were prepared 24 hours in advance at the experimental temperature, have been studied batch wise at 50&ordm / C. Glucose adsorption, in solutions that had adsorption differences, was fast on both zeolites, on the contrary of slow adsorption of fructose with the stable dynamics. Both adsorptions had small amounts of adsorption changes after minute 30. The treatments made under the same conditions with the same mixtures showed Ca-Y zeolite had better separation capacity compared to H-Y zeolite. Some trials were repeated with CaCl2 added to the solutions. The slowed down affection of fructose adsorption in spite of the small change of glucose adsorption led to better separation. Samples were analyzed by classical methods, not HPLC. All the data were considered with various models and their convergence numbers were tested for their closeness to reality. The models were analyzed by response surface methodology and some of those models had correlation factors as high as 88% at the equilibrium points at 30th minutes. Besides, time dependent models have been considering the lag times with a time dependent variable included all the data of all treated solutions with correlation as high as 79.5%.

Reproduction de : Thèse de doctorat : Sciences de la Vie et de la Santé : Lille 1 : 2004. Reproduction de : Thèse de doctorat : Sciences de la Vie et de la Santé : Universitatea "Al. I. Cuza", Iasi : 2004. / Thèse en cotutelle. N° d'ordre (Lille 1) : 3479. Titre provenant de la page de titre du document numérisé. Bibliogr. p. 196-223.

Thesis (M. Phil.)--University of Hong Kong, 1972. / Typewritten.

Standards of Amadori rearrangement products (ARP) were synthesized for the purpose of developing analytical techniques and performing mechanistic studies related to their thermal decomposition. Several synthetic strategies were explored. An HPLC analytical system with a diode array detector was coupled to a fluorometer and an electrochemical detector, in order to detect simultaneously and on-line, a wide variety of degradation products of ARPs and to follow their kinetics. The potential of such a system to analyze complex Maillard mixtures was demonstrated. The kinetics of the reaction of glucose with morpholine (a Strecker inactive analogue of proline was used in order to simplify the kinetics) to produce Amadori morpholine was studied under experimental conditions that minimize side reactions and maximize Amadori product formation. At specific time intervals, the samples were analyzed for the presence of reactants and Amadori product by the multidetector HPLC system. Color and fluorescence were also measured. The data obtained were used to calculate the rate constants for the formation and degradation of Amadori product. A mechanistic model that statistically fitted the kinetic data was proposed. To further understand the details of the decomposition mechanism of Amadori proline, different mass spectrometric experiment were performed. High resolution, linked-field scan and neutral loss experiments have indicated that 1-((2$ sp prime$-carboxyl)pyrrolidinyl)-1-deoxy- scD-fructose (Proline Amadori product) followed two main pathways of fragmentation under electron impact conditions; one initiated by the ring oxygen and the other by the amino acid nitrogen, producing two well stabilized fragment ions; oxonium and imminium ions. In addition, ortho-elimination reactions initiated by O or N-centered radical sites were shown to produce the most intense peaks and diagnostically important ions for the identification of Amadori products. However this approach can only pro

No description available.

High chronic fructose consumption has been linked to many diseases. Sugar metabolites, especially glyoxal and methylglyoxal can form advanced glycation products, which contribute to the pathology of diabetic complications. Our objective was to study the metabolism of these metabolites and the associated protein carbonyation and cytotoxicity in isolated hepatocytes. In addition, the effect of oxidative stress on the metabolism of these toxins was also investigated. Methylglyoxal and glyoxal can induce protein carbonylation, which contributes to hepatocyte toxicity. Methylglyoxal, but not glyoxal, was detoxified mainly by the glyoxalase system. Both toxins can be metabolized by mitochondrial aldehyde dehydrogenase. The detoxification of glyoxal was impaired under oxidative stress conditions (i.e. increased hydrogen peroxide level). Glyoxal was found to be a common autoxidation product from glyceraldehyde, hydroxypyruvate and glycolaldehyde. Glyoxal and the reactive oxygen species formation during the autoxidation process contributed to the hepatocyte toxicity of glyceraldehyde, hydroxypyruvate and glycolaldehyde.

High chronic fructose consumption has been linked to many diseases. Sugar metabolites, especially glyoxal and methylglyoxal can form advanced glycation products, which contribute to the pathology of diabetic complications. Our objective was to study the metabolism of these metabolites and the associated protein carbonyation and cytotoxicity in isolated hepatocytes. In addition, the effect of oxidative stress on the metabolism of these toxins was also investigated. Methylglyoxal and glyoxal can induce protein carbonylation, which contributes to hepatocyte toxicity. Methylglyoxal, but not glyoxal, was detoxified mainly by the glyoxalase system. Both toxins can be metabolized by mitochondrial aldehyde dehydrogenase. The detoxification of glyoxal was impaired under oxidative stress conditions (i.e. increased hydrogen peroxide level). Glyoxal was found to be a common autoxidation product from glyceraldehyde, hydroxypyruvate and glycolaldehyde. Glyoxal and the reactive oxygen species formation during the autoxidation process contributed to the hepatocyte toxicity of glyceraldehyde, hydroxypyruvate and glycolaldehyde.

Mutationen im Gen der Aldolase B auf Chromosom 9 führen zur „hereditären“ Fructoseintoleranz (HFI), die nach einer längeren Fructoseexposition zu schweren Leber- und Nierenschäden bis hin zum völligen Versagen dieser Organe führen kann. Zum Ausschluß bzw. zur Bestätigung einer HFI werden seit ca. 20 Jahren hauptsächlich molekularbiologische Verfahren eingesetzt. Dabei zeigte es sich, dass wenige, regional unterschiedlich häufig vorkommende Mutationen (in Deutschland A149P und A174D) bei dem Großteil der Patienten für diese Erkrankung verantwortlich sind. Viele Laboratorien beschränken sich daher auf den Nachweis bzw. den Ausschluß dieser Mutationen. Ziel der vorliegenden Arbeit war es zu zeigen, dass auch bei den Patienten mit der Verdachtsdiagnose „hereditäre Fruktoseintoleranz“, bei denen die häufig vorkommen-den Mutationen A149P und A174D nur auf einem oder auf keinem der zwei Allele nachweisbar waren, die Diagnose mit molekularbiologischen Methoden durch den Nachweis seltener oder bisher nicht beschriebener Mutationen gesichert werden kann. Untersucht wurden 8 Patienten aus 5 Familien mit dieser Verdachtsdiagnose. Dabei handelte sich um Patienten mit klinischen Symptomen und in einem Fall enzymatisch gesicherter Diagnose. In allen Fällen mit Ausnahme von Patient III3 (Genotyp A174T/Wildtyp) konnten zwei Mutationen im Aldolase B-Gen nachgewiesen werden. Von den insgesamt identifizierten 8 Mutationen waren 3 zum Zeitpunkt der Durchführung der experimentellen Arbeiten in der Fachliteratur noch nicht beschrieben und erhöhen die Zahl der bekannten Mutationen im Aldolase B – Gen auf insgesamt 45.