| Summary: | Dissertação de mestrado em Bioquímica Aplicada (especialização em Biomedicina) Enzymes are biological catalysts that accelerate the rate of chemical reactions in cells and are commonly used in a range of industries. They can improve the efficiency, cost-effectiveness and environmental impact of many processes and enhance the characteristics and quality of products. New enzymes with novel backbone sequences and novel functions and characteristics are constantly called for. In the present project, 116 yeasts from mainly acidic pH biowastes, 11 aquatic filamentous fungi from cold environments in Portugal and Iceland, 3 bacterial isolates and 3 metagenomic clones from Irish soil samples were screened for the production of glycoside hydrolase enzymes of industrial interest, namely xylanases, cellulases, pectinases and chitinases. Methods were investigated and developed for a simplified screening of the various isolates at 16 ºC, pHs 4, 7 and 9, with, in particular, the development of a novel xylanase substrate. The substrate was shown to be suitable for measurement of xylanase activity with the commonly used DNS assay but further tests with other xylanases are recommended to confirm this. Furthermore, the substrate was successfully coupled with remazol brilliant blue (RBB) dye, obtaining a chromogenic substrate and enabling a highly sensitive and efficient plate based screening of xylanases. Screening of the CBMA culture collection isolates allowed for identification of a large number of glycoside hydrolase positive isolates, with 31 isolates showing desired activities exclusively at acidic pH. These enzymes may be of interest for use in various applications and, in particular, in food and beverages applications. In addition, 5 isolates were found to display all 4 activities screened for at pH 4, and these isolates may have potential for use in biomass treatment applications. A cellulase positive metagenomic clone with metagenomic DNA from Irish soil was selected for further analysis. The gene for this cellulase ...
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