Isolation of cytoplasmic NADPH-dependent phenol hydroxylase and catechol-1,2-dioxygenase from Candida tropicalis yeast

The efficiencies of NADPH-dependent phenol hydroxylase (EC 1.14.13.7) and catechol 1,2-dioxygenase (EC.1.13.11.1) in biodegradation of phenol in the cytosolic fraction isolated from yeast Candida tropicalis were investigated. Enzymatic activities of both NADPH-dependent phenol hydroxylase and catechol 1,2-dioxygenase were detected in the cytosolic fraction of C. tropicalis grown on medium containing phenol. Using the procedure consisting of chromatography on DEAE-Sepharose, fractionation by polyethylene glycol 6000 and gel permeation chromatography on Sepharose 4B the enzyme responsible for phenol hydroxylation in cytosol, NADPH-dependent phenol hydroxylase, was isolated from the cytosolic fraction of C. tropicalis close to homogeneity. However, fractionation with polyethylene glycol 6000 lead to a decrease in catechol 1,2-dioxygenase activity. Therefore, another procedure was tested to purify this enzyme. Gel permeation chromatography of proteins of the eluate obtained by chromatography on a DEAE-Sepharose column was utilized to separate phenol hydroxylase and catechol 1,2-dioxygenase. Among gel permeation chromatography on columns of Sephadex G-100, Sephacryl S-300 and Sepharose 4B tested for their efficiencies to isolate phenol hydroxylase and catechol 1,2-dioxygenase, that on Sephacryl S-300 was found to be suitable for such a procedure. Nevertheless, even this chromatographic method did not lead to obtain catechol 1,2-dioxygenase in sufficient amounts and purity for its further characterization. The data demonstrate the progress in resolving the enzymes responsible for the first two steps of phenol degradation by the C. tropicalis strain

Novověké polní fortifikace – možnosti studia a jejich interpretace

The thesis is focused on documentation and identifi cation of postmedieval field fortifications. Fortifications were searched on the sheets of 1st Military Survey. There were collected 60 objects in different state of disintegration. Several analyses in GIS were aimed to fi nd out and verify the relations of the fortifi cations to the terrain. Elevation, proximity to watercourse and roads were detected. Based on the combination of diff erent sources the fortifications were dated to the War of Bavarian heritage (1778–1779), though it might be said that their strategic positions have been used in previous conflicts as well. The paper tried to determine the possibilities of cartographic sources and how much information they provide. The final conclusion is that such sources are ideal for studying the postmedieval/modern landscape. Research of the remains of field fortifications should be done with the usage of various sources. Proper research is based on the knowledge of possible written sources followed by iconography, toponomastic, cartographic and especially archaeological sources which, used in combination, aim to the best results of the research