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Biochemical characterization of polygalacturonases from five different isolates of Botrytis cinerea

TitoloBiochemical characterization of polygalacturonases from five different isolates of Botrytis cinerea
Tipo di pubblicazioneArticolo su Rivista peer-reviewed
Anno di Pubblicazione2002
AutoriPashkoulov, D., Giannetti I., Benvenuto Eugenio, and De Martinis D.
RivistaMycological Research
Volume106
Paginazione827 - 831
Data di pubblicazione2002
ISBN Number09537562 (ISSN)
Parole chiavebiochemical composition, Botrytis cinerea, fungal disease
Abstract

Polygalacturonase (PG) secretion was analysed from five independent isolates of Botrytis cinerea grown in minimal liquid media enriched with pectin as the only carbon source. To acquire information about the biochemical characteristics of the PGs secreted in vitro, we combined enzyme-activity assays with Western blot analysis and isoelectro-focusing (IEF). All isolates secreted at least four PGs isozymes with a similar electrophoretic patterns, easily distinguishable by western blots. IEF enabled the isoelectric point of the PGs for each isolate to be determined. Wide variability in isoelectric point was observed between the PGs of each isolate, ranging from above pH 9.0 to below pH 5.0. For one of the isolates, each PG secreted in vitro was further purified by chromatofocusing and analysed for endo/eso-polygalacturonase activity by thin layer chromatography. This combination of analytical techniques and enzyme assays provided a view of the pool of PGs produced by B. cinerea and defined the biochemical features of each isozyme.

Note

Cited By :4Export Date: 16 July 2015CODEN: MYCRECorrespondence Address: De Martinis, D.; ENEA, UTS Biotecnologie, Sezione Genetica e Genomica Vegetale, C.R. Casaccia, Via Anguillarese 301, C.P. 2400, 00060 Roma, Italy; email: demartinis@casaccia.enea.itReferences: Adrian, M., Rajaei, H., Jeandet, P., Veneau, J., Bessis, R., Resveratrol oxidation in Botrytis cinerea conidia (1998) Phytopathology, 88, pp. 472-476;Alghisi, P., Favaron, F., Pectin-degrading enzymes and plant-parasite interactions (1995) European Journal of Plant Pathology, 101, pp. 365-375; Barkai-Golan, R., Lavy-Meir, G., Kopeliovich, E., Pectolytic and cellulolytic activity of Botrytis cinerea Pers. related to infection of non-ripening tomato mutants (1988) Journal of Phytopathology, 123, pp. 174-183; Benito, E.P., Ten Have, A., Van’t Klooster, J.W., Van Kan, J.A.L., Fungal and plant gene expression during synchronized infection of tomato leaves by Botrytis cinerea (1998) European Journal of Plant Pathology, 104, pp. 207-220; Cruickshank, R.H., Wade, G.C., Detection of pectin enzymes in pectin-acrilamide gels (1980) Analytical Biochemistry, 107, pp. 177-181; Fernandez, N., Patino, B., Vazquez, C., Pectin degrading enzymes secreted by six isolates of Fusarium oxysporum (1993) Mycological Research, 97, pp. 461-466; Jarvis, W., Botrytinia and Botrytis species: Taxonomy and pathogenicity (1977) Canada Department of Agriculture Monograph No. 15, , Agriculture Canada, Ottawa; Johnston, D.J., Williamson, B., Purification and characterisation of four polygalacturonases from Botrytis cinerea (1992) Mycological Research, 96, pp. 343-349; Johnston, D.J., Williamson, B., An immunological study of the induction of Polygalacturonases in Botrytis cinerea (1992) FEMS Microbiology Letters, 20, pp. 19-24; Lang, C., Dornenburg, H., Perspectives in the biological function and the technological application of polygalacturonases (2000) Applied Microbiology and Biotechnology, 53, pp. 366-375; Leone, G., Schoffelmeer, E.A.M., Van den Heuvel, J., Purification and characterisation of a constitutive polygalacturonase associated with the infection process of french beans leaves by Botrytis cinerea (1990) Canadian Journal of Botany, 68, pp. 1921-1930; Lever, M., A new reaction for colorimetric determination of carbohydrates (1972) Analytical Biochemistry, 47, pp. 273-279; Mansfield, J.W., Mechanism of resistance to Botrytis (1980) The Biology of Botrytis, pp. 181-218. , (J. R. Coley-Smith, K. Verhoeff & W. R. Jarvis, eds). Academic Press, New York; Robertson, E.F., Dannelly, K., Malloy, P.J., Reeves, H., Rapid isoelectric focusing in a vertical polyacrilamide minigel system (1987) Anals of Biochemistry, 167, pp. 290-294; Ten Have, A., Breuil, W.O., Wubben, J.P., Visser, J., Van Kann, J.A., Botrytis cinerea endopolygalacturonase genes are differentially expressed in various plant tissues (2001) Fungal Genetics and Biology, 33, pp. 97-105; Ten Have, A., Mulder, W., Visser, J., Van Kan, J.A.L., The endopolygalacturonase gene Bcpg1 is required for full virulence of Botrytis cinerea (1998) MPMI, 11, pp. 1009-1016; Urbanek, H., Zalewska-Sobczak, J., Multiplicity of cell wall degrading glycosidic hydrolases produced by apple infecting Botrytis cinerea (1984) Phytopatologische Zeitschrift, 110, pp. 261-271; Van der Cruyssen, G., De Meester, E., Kamoen, O., Expression of polygalacturonases of Botrytis cinerea in vitro and in vivo (1994) Mededelingen van de Facultent Landbouw Wetonshappen Rijksuniversitert Gent, 59, pp. 895-905; Verhoeff, K., Warren, J.M., In vitro and in vivo production of cell wall degrading enzymes by Botrytis cinerea from tomato (1972) Netherlands Journal of Plant Pathology, 78, pp. 179-185; Wubben, J.P., Mulder, W., Ten Have, A., Van Kan, J.A.L., Visser, J., Cloning and partial characterisation of endopolygalacturonase genes from Botrytis cinerea (1999) Applied and Environmental Microbiology, 20, pp. 1596-1602; Wubben, J.P., Ten Have, A., Van Kan, J.A.L., Visser, J., Regulation of endopolygalacturonase genes expression in Botrytis cinerea by galacturonic acid, ambient pH and carbon catabolite repression (2000) Current Genetics, 37, pp. 152-157

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