Universität Tübingen Botanisches Institut
Lehrstuhl Physiologische Ökologie der Pflanzen
Wisser G, Guttenberger M, Hampp R, Nehls U (2000) Identification and characterization of an extracellular acid trehalase from the ectomycorrhizal fungus Amanita muscaria. New Phytologist (in press) The non-reducing disaccharide trehalose is a major storage compound in most fungi. For a better understanding of carbon metabolism in the ectomycorrhizal-forming basidiomycete A. muscaria, trehalase activity was analyzed from mycelium growing in liquide culture or on agar plates, and from mycorrhizas. Trehalase activity was found in both the culture medium and in mycelial extracts. The excreted trehalase was purified to apparent homogeneity by anion-exchange chromatography, gel-filtration and preparative gel-electrophoresis. The identified enzyme belongs to the group of acid trehalases. The apparent molecular mass of the native enzyme was estimated to be 165 kDa by gel-filtration and 135 kDa by SDS-PAGE. Isoelectrofocusing indicated a pI of about 3.7, which is typical for acid trehalases. The enzyme is highly specific for its substrate trehalose with an apparent Km value of 0.38 mM. Validamycin and sucrose act as competitive inhibitors with Ki values of 45 nM and 15 mM, respectively. The activity of acid trehalase which is excreted into the growth medium, is independent of the carbon source (glucose or trehalose) revealing that the enzyme is not regulated by its substrate trehalose. Nevertheless, fungal hyphae grown in the absence of an external carbon source showed enhanced enzyme excretion. The biochemical characteristics of the trehalase activity meseared in the mycelium are in the same range as those determined for the excreted one. The enzyme is localised in the cell wall and its activity is strongly reduced in ectomycorrhizas.
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