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Avenacosidase from oat: purification, sequence analysis and biochemical characterization of a new member of the BGA family of β-glucosidases

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Abstract

A protein consisting of 60 kDa subunits (As-P60) was isolated from etiolated oat seedlings (Avena sativa L.) and characterized as avenacosidase, a β-glucosidase that belongs to a preformed defence system of oat against fungal infection. The enzyme is highly aggregated; it consists of 300–350 kDa aggregates and multimers thereof. Dissociation by freezing/thawing leads to complete loss of enzyme activity. The specificity of the enzyme was investigated with para-nitrophenyl derivatives which serve as substrates, in decreasing order β-fucoside, β-glucoside, β-galactoside, β-xyloside. The corresponding orthonitrophenyl glycosides are less well accepted. No hydrolysis was found with α-glycosides and β-thioglucoside. An anti-As-P60 antiserum was prepared and used for isolation of a cDNA clone coding for As-P60. A presequence of 55 amino acid residues was deduced from comparison of the cDNA sequence with the N-terminal sequence determined by Edman degradation of the mature protein. The presequence has the characteristics of a stroma-directing signal peptide; localization of As-P60 in plastids of oat seedlings was confirmed by western blotting. The amino acid sequence revealed significant homology (>39% sequence identity) to β-glucosidases that are constituents of a defence mechanism in dicotyledonous plants. 34% sequence identity was even found with mammalian and bacterial β-glucosidases of the BGA family. Avenacosidase extends the occurrence of this family of β-glucosidases to monocotyledonous plants.

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Authors and Affiliations

  1. Botanisches Institut, Universität München, Menzinger Strasse 67, D-80638, München, Germany

    Sabine Gus-Mayer, Harald Brunner & Wolfhart Rüdiger

  2. Botanisches Institut der Universität zu Köln, Gyrhofstrasse 15, D-50931, Köln, Germany

    Hansjörg A. W. Schneider-Poetsch

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  1. Sabine Gus-Mayer
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  2. Harald Brunner
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  3. Hansjörg A. W. Schneider-Poetsch
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  4. Wolfhart Rüdiger
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Gus-Mayer, S., Brunner, H., Schneider-Poetsch, H.A.W. et al. Avenacosidase from oat: purification, sequence analysis and biochemical characterization of a new member of the BGA family of β-glucosidases. Plant Mol Biol 26, 909–921 (1994). https://doi.org/10.1007/BF00028858

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