Thesis (M.S.)--Virginia Polytechnic Institute and State University, 1992. / Vita. Abstract. Includes bibliographical references (leaves 46-49). Also available via the Internet.

Göttingen, Univ., Diss., 2004. / Computerdatei im Fernzugriff.

Göttingen, Universiẗat, Diss., 2004.

Submitted by Biblioteca da Faculdade de Farmácia (bff@ndc.uff.br) on 2017-04-19T16:57:00Z No. of bitstreams: 1 Pontes, Simone Gomes [Dissertação, 2012].pdf: 1302242 bytes, checksum: 7aab3cecd1b771d41f103c491d846579 (MD5) / Made available in DSpace on 2017-04-19T16:57:00Z (GMT). No. of bitstreams: 1 Pontes, Simone Gomes [Dissertação, 2012].pdf: 1302242 bytes, checksum: 7aab3cecd1b771d41f103c491d846579 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A dihidroxiacetona (DHA) é uma molécula constituída por três carbonos e não tóxica, utilizada como insumo para as indústrias de cosméticos, fármacos e química fina. É produzida industrialmente por fermentação, utilizando a bactéria Gluconobacter oxydans. Esse processo tem como principal limitação a inibição do crescimento tanto pelo substrato – glicerol – quanto pelo produto – DHA e, por tal, estudos recentes descrevem propostas para melhoria do processo. Sendo a conversão de glicerol a DHA realizada por uma única enzima em uma etapa, o presente trabalho considera que tal processo se enquadra nas definições de uma biotransformação, ou seja, a utilização de um catalisador biológico com o propósito de converter um substrato a um produto estruturalmente similar, através de modificações específicas e utilizando um número limitado de etapas enzimáticas. Dessa forma, neste estudo foram avaliados comparativamente a secagem de células em acetona e, em um segundo momento, a utilização de células de Gluconobacter oxydans previamente crescidas, para a produção de DHA a partir de glicerol. Objetivando contornar o principal problema do processo, que é a inibição do crescimento microbiano pelo substrato e pelo produto, foram testadas duas linhagens. A utilização de células secas em acetona se mostrou possível, porém os resultados não foram reprodutíveis e células previamente crescidas por 24 horas passaram a ser usadas nos experimentos de biotransformação. O pH e a temperatura de reação foram selecionados a partir de um planejamento delineamento composto central rotacional como sendo de 34ºC e pH de 4,5, para G. oxydans CCT 0552 e de 26ºC e pH de 4,5 para G. oxydans CCT 0174. A linhagem G. oxydans CCT 0552 se mostrou mais adequada à oxidação de glicerol à DHA, com aumento do acúmulo de DHA no meio reacional com o tempo (2,1 g/g biomassa) e com a produtividade constante (0,45 g/g biomassa). Foi constatada perda de atividade nas células estocadas por congelamento, o que leva à necessidade de selecionar um melhor método de conservação das células para a utilização na produção / The dihydroxyacetone (DHA) is a non-toxic molecule consisting of three carbons, used in the cosmetics, pharmaceuticals and fine chemicals industry. The DHA is industrially produced by fermentation, using the bacteria Gluconobacter oxydans. The main bottleneck of this process is the growth inhibition by the substrate – glycerol – and the product – DHA. This problem leads recent studies to describe proposals for improving the process. As the conversion of glycerol to DHA is performed by a single enzyme in one step, this study considers that this process fits in the definitions of biotransformation, in other words, the use of a biological catalyst in order to convert a substrate for a structurally similar products, by speficic modifications, and using a limited number of enzymatic steps. Thus, this study were assessed by comparison with drying of cells in acetone and in second stage, the use of previously grown cells of Gluconobacter oxydans for the production of DHA from glycerol. The use of dried cells proved to be possible, but the results were not reproducible and the biotransformation experiments were done with previously grown cells of 24 hours age. . The best pH and temperature for the reaction were selected from a central composite design as being 34o C and pH 4.5 for G. oxydans CCT 0552 and 26o C and pH 4.5 for G. oxydans CCT 0174. The strain G. oxydans CCT 0552 was more suitable for the oxidation of glycerol to DHA, with increased accumulation of DHA in the reaction media (2,1 g/g biomass) and constant productivity (0,45 g/g biomass). Loss of activity was observed in cells stored by freezing, which leads to the need to select a best method of preserving cells for the production

Thesis (Ph. D.)--University of Wisconsin. 1965. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Techn. Universiẗat, Diss., 2005--München.

M.S.

The gluconobacters are well known for their plasma membrane-bound dehydrogenases that rapidly oxidize compounds and release the products into the medium. In 1989, Qazi et al. proposed that genes coding for membranebound glucose dehydrogenase are on a plasmid in G. oxydans strain ATCC 9937. The only other known report of gluconobacter plasmids was by Fukaya, et al. who reported that 23 of 36 strains examined contained plasmids. I wish to learn more about the presence and significance of plasmids in the gluconobacters. In this study, I selected 14 strains representing the three gluconobacter species to determine possible similarities and differences in their plasmid profiles. To date, our most successful method for extracting plasmids involves alkaline lysis of cells at 60°C followed by phenol-chloroform extraction which leaves the nucleic acids in the aqueous phase. These nucleic acids are then precipitated in a LiCl-ethanol solution using glycogen as a carrier, and plasmids are separated on agarose gels. We found that 11 of the 14 gluconobacter strains surveyed contain plasmids ranging in size from 2.7 to greater than 200 kb. The type strain of the genus (G. oxydans strain ATCC 19357) contained 6 plasmids ranging in size from 4 to 120 kb. Strains from G. frateurii and G. asaii also contained a wide range of plasmid sizes. Hybridization techniques were utilized to determine if plasmids of similar size are genetically similar. The 2.7 and 16.2 kb plasmids of G. oxydans strain ATCC 621 and IFO 12528 were shown to share homology. The 100 and 120 kb plasmids of G. oxydans strain ATCC 19357 hybridized with the 250 kb plasmids of ATCC 621 and the 210 kb plasmid of ATCC 9937. Also showing homology were the 6.4 kb plasmids of G. oxydans strains ATCC 9937 and IFO 12467. / Master of Science

Bacteria of the genus Gluconobacter have very active, membrane-bound, NAD(P)-independent, polyol dehydrogenases which stoichiometrically produce the singlestep oxidation product of polyols provided in the growth medium. These bacteria have a high respiratory quotient which is believed to result from oxidations by these dehydrogenases. These organisms grow and survive at pH values as low as 2.5 leading to speculation that their membrane-bound dehydrogenase activity provides the rapid electron flow necessary to purge cells of toxic levels of hydrogen ions. These dehydrogenases are also believed to be used for energy metabolism, and there is no clear understanding of their function in the cell metabolism. Oxidation of sorbitol in Gluconobacter oxydans ATTC 621 was studied to determine if the oxidations by the membrane bound sorbitol dehydrogenase (mSDH) were required for growth, and whether they functioned to protect the cells in low pH environments. G. oxydans required a high concentration of sorbitol in the medium, and a reduction in the concentration to 0.1% decreased the rate and extent of growth. Using mutants with decreased levels of mSDH, we found that growth rates decreased as a result of this mutation, indicating that mSDH activity was needed for growth. No changes in the specific activity of mSDH in strain ATCC 621 occurred when the cells were grown at pH 7.0, 6.0, and 4.5. However, cytochrome levels were doubled in cells grown at pH 4.5 compared to pH 6.0 and 7.0. The increased cytochrome levels did not increase the oxygen uptake of the pH 4.5 grown cells on sorbitol. Cells grown at all pH values respired more rapidly when tested at pH 4.5, and respiration decreased as the pH increased. The higher activity at lower pH values may result from increased efficiency of mSDH, which has an in vitro pH optimum of 5.2. Magnesium and calcium increased the respiration of pH 6.0 grown cells but not pH 4.5 grown cells. Less cell mass per mg of sorbitol oxidized was obtained when cells were grown at pH 4.5 compared to pH 6.0 and 7.0. However, no differences were detected in the specific activity of any of the sorbitol oxidizing enzymes. The activity of mSDH in G. oxydans is necessary for the growth of this bacterium. The mSDH specific activity is not regulated by the growth pH, but increased levels of cytochromes and decreased cell yields indicate a change in the cell's oxidative system resulting from lowered growth pH values. / Ph. D.

Thesis (doctoral)--Universität Kiel, 2000.