Candida tropicalis yeast and bacteria Comamonas testosteroni have been considered to be able to metabolize phenol and utilize it as the only source of carbon and energy. In our laboratory we investigated the cytoplasmic enzymes responsible for the first and second step of phenol degradation, NADPH-dependent phenol hydroxylase of both C. tropicalis and C. testosteroni and catechol 1,2-dioxygenase of C. tropicalis. The aim of our study was to isolate and partially characterize those enzymes. Phenol hydroxylase purification consisted of preparation of cytosol from C. tropicalis yeast by fraction centrifugation, chromatography and re-chromatography on a column of DEAE Sepharose, fractionation by precipitation of the enzyme with polyethylene glycol 6000 and gel permeation chromatography on a column of Sephacryl S-300. Extracellular phenol hydroxylase of C. testosteroni was purified by fraction precipitation with polyethylene glycol 6000 and by gel permeation chromatography on 4B Sepharose and Sephacryl S-300. Catechol 1,2-dioxygenase was purified using the procedure consisting of: chromatography and re- chromatography on a column of DEAE Sepharose, lyophilization of the enzyme and gel permeation chromatography on a column of Sephadex G-100. The enzyme activity was determined by two methods: use of HPLC...
| Identifer | oai:union.ndltd.org:nusl.cz/oai:invenio.nusl.cz:311550 |
| Date | January 2011 |
| Creators | Vilímková, Lenka |
| Contributors | Stiborová, Marie, Entlicher, Gustav, Mareš, Jaroslav |
| Source Sets | Czech ETDs |
| Language | Czech |
| Detected Language | English |
| Type | info:eu-repo/semantics/doctoralThesis |
| Rights | info:eu-repo/semantics/restrictedAccess |
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