Purification and properties of two enzymes from Dichomitus squalens which exhibit both cellobiohydrolase and xylanase activity |
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| Institution: | 1. Department of Analytical Chemistry, Institute of Chemistry of Araraquara, São Paulo State University (UNESP), CP 355, 14801-970 Araraquara, São Paulo, Brazil;2. Center for Monitoring and Research of the Quality of Fuels, Biofuels, Crude Oil and Derivatives (CEMPEQC), Institute of Chemistry of Araraquara, São Paulo State University (UNESP), CP 355, 14801-970 Araraquara, São Paulo, Brazil;3. Institute of Research on Bioenergy (IPBEN), São Paulo State University (UNESP), 13500-230 Rio Claro, São Paulo, Brazil;1. Institute of Microbiology, Faculty of Biology, University of Warsaw, Miecznikowa 1, 02-096 Warsaw, Poland;2. RDLS Ltd., Miecznikowa 1, 02-096 Warsaw, Poland;3. Polish Geological Institute - National Research Institute, Rakowiecka 4, 00-975, Warsaw, Poland;4. Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-096, Warsaw, Poland;1. Laboratorio de Biotecnología Molecular, Instituto de Biotecnología Misiones, CONICET, Facultad de Ciencias Exactas Químicas y Naturales, Universidad Nacional de Misiones, CP3300, Posadas, Misiones, Argentina;2. Laboratorio de Micología Experimental, Dpto. de Biodiversidad y Biología Experimental, FCEN, UBA, INMIBO (CONICET), 1428, CABA, Argentina |
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| Abstract: | Two fractions (Ex I and II), exhibiting activity towards p-nitrophenyl β-cellobioside (pNPC) have been isolated by chromatofocusing of the proteins obtained from the supernatant solution of a cellulose-containing culture of the white-rot fungus Dichomitus squalens. They were further purified up to 16.0- and 14.2-fold by chromatography on Phenyl-Sepharose CL-4B and ion-exchange chromatography on DEAE-Trisacryl. Each purified enzyme gave a single peak for protein and activity on chromatography on Ultrogel AcA-54 and a single protein band in disc gel electrophoresis, in the absence and presence of sodium dodecyl sulphate, and on isoelectrofocusing. The mol. wts. of Ex I and II were 39,000 and 36,000, respectively, and their isoelectric points were 4.6 and 4.5, respectively. Maximum activity towards pNPC was shown at pH 5.0 and 60°, and each enzyme was stable over the pH range 4.0–8.0, and up to 70° and 60° for Ex I and II, respectively. The enzymes cleaved pNPC, released mainly cellobiose from cellulose, were especially active towards xylan and o-nitrophenyl β-d-xylopyranoside, and exhibited strong transglycosylating activities. |
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