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Báo cáo lâm nghiệp: "Production and characterization of exocellular in ectomycorrhizal fungi proteases"

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Tuyển tập các báo cáo nghiên cứu về lâm nghiệp được đăng trên tạp chí lâm nghiệp Original article đề tài: Production and characterization of exocellular in ectomycorrhizal fungi proteases...

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Production and characterization of exocellular proteases in ectomycorrhizal fungi B. Botton K. El-Badaoui Universitc de Nancy I, Facult6 des Sciences, Laboratoire de Physiologie V6g6tale et Foresti6re, BP 239, 54506 Vand&oelig;uvre-Iès-Nancy Cedex, France serum albumin (BSA), gelatin, casein or pro- Introduction teins extracted from the litter (Botton et al., 1986). Enzyme activities were determined in Mycorrhizas grow very often in soil layers culture filtrates, either by spectrophotometry or in which organic nitrogen compounds are by spectrofluorometry using fluorescein isothio- cyanate-labeled BSA (FITC-BSA) as described present in large quantities. The ability of by Twining (1984). Proteins purified from the several ectomycorrhizal fungi to assimilate litter by ammonium sulfate fractionation and proteins and to transfer their nitrogen to DEAE-cellulose chromatography were also plants has already been demonstrated labeled with FIT’C and used as a substrate and (Abuzinadah and Read, 1986; Abuzinadah as inducers. al., 1986). These results suggest that et proteases are excreted by the fungi and Results protease activities detected in mycorrhiza- forming fungi were reported by Lyr (1963) Protease activity remained at a very low and Ramstedt and S6derhall (1983). In level when the fungus was cultivated in this study, the process of production of the presence of ammonium, but increased exocellular proteases has been investi- strongly after transfer of the organism to gated in free living fungi as well as in fresh medium containing casein, gelatin or mycorrhizas. Experiments were carried BSA as the sole nitrogen source. Gelatin out with Amanita rubescens and Lacta- proved to be the most efficient inducer rius subdulcis which live in organic hori- (Fig. 1a). Elimination of carbon, inorganic zons and with Cenococcum geophilum nitrogen or sulfur was not sufficient to and Hebeloma crustuliniforme which live induce exoceillular proteases, proteins predominantly in mineral soil layers. also had to be present. However, in nitrogen- and carbon-deficient media, gelatin was not efficient (not shown). A Materials and Methods group of proteins of about 46 kDa was purified from the forest litter and was a The were collected from mycorrhizas. fungi better inducer than gelatin, in spite of its grown in shaken cultures at 25°C in They were lower concentration, both in C. geophilum Pachlewski’s medium containing nitrogen as and H. crustuliniforme (Fig. 1 b). either diammonium tartrate or proteins: bovine
As shown in Table I, A. rubescens and L. subdulcis secreted larger amounts of exocellular proteases than C. geophilum and H. crustuliniforme, although growth of the 2 former fungi was considerably reduced. For the 4 fungi tested, controls with ammonium (Pachlewski’s medium) were better developed than the cultures grown in the presence of proteins from the litter, but protease excretion was reduced. Protease activity was also induced in beech-Lactarius ectomycorrhizas incubat- ed in the presence of litter proteins. Gelatin was less efficient and ammonium repressed the excretion after 48 h of incubation. Non-mycorrhizal roots did not exhibit protease activity (Fig. 2). Conclusion These results indicate that ectomycor- with proteins. Very often, supplemented detected in the rhizal amounts of alkaline fungi significant secrete proteases were medium; however, when the fungi proteases when the culture medium is were
between the ability of the fungi to produce such enzymes and their distribution in the soil layers. References Abuzinadah R.A. & Read D. (1986) The role of proteins in the nitrogen nutrition of ectomycor- rhizal plants. I. Utilization of peptides and proteins by ectomycorrhizal fungi. New Phytol. 103, 481-493 Abuzinadah R.,A., Finlay R.D. & Read D.J. (1986) The role of proteins in the nitrogen nutrition of ectomycorrhizal plants. 11. Utilization of proteins by mycorrhizal plants of Pinus contorta. New F’hytoG 103, 495-506 Botton B., El-Badaoul K. & Martin F. (1986) Induction of extracellularproteases in the ascomycete Cenococcum geophilum. In: Phy- siological and Genetical Aspects of Mycor- rhizae. Proceedings of the Ist European Symposium on Mycorrhizae, Dijon, July 1985. INRA, Paris, pp. 403-406 Lyr H. (1963) IVlykorrhiza. In: Infernationales Mykorrhizazymposium. Jena, 1963. Fisher, Verlag, pp. 123-145 induced with proteins from the litter and Ramstedt M. & S6derhall K. (1983) Protease, assayed with the same proteases were phenoloxidase and pectinase activities in substrate, optimum pH of activity was mycorrhizal fungi. Trans. Br. Mycol. Soc. 81, about 5. It is likely that the proteases 157-161 excreted enable the organism to use Twining S.S. (1984)Fluorescein isothiocyanate- extracellular proteins as a source of labeled casein assay for proteolytic enzymes. nitrogen and a close correlation exists Anal. Biochem. 143, 30-34

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