Higher fungi are a rich source of L-amino acid oxidases

Abstract

L-Amino acid oxidases (LAO) are widely distributed enzymes but those from snake venoms have been studied the most. We describe a method for in-gel detection of LAO activities based on H₂O₂ detection by a horseradish peroxidase-coupled reaction using o-phenylenediamine. Complex substrates and single L-amino acids were used successfully for screening LAO activities in higher fungi using crude aqueous extracts of fruiting bodies of 22 basidiomycetes and 1 ascomycete. Half of these samples exhibited one to two bands of LAO activities with mostly broad substrate specificities and a variety of apparent molecular masses ranging from 25 to 200 kDa that were generally more active at pH 5.5 than at pH 8.0. Mushrooms are shown to be a rich source of LAOs that could find use in various medical and biotechnological applications.

Keywords: Fruiting body; Fungi; Horseradish peroxidase; In-gel detection; L-Amino acid oxidase; Mushroom.

Fig. 1

In-gel LAO activity at pH 5.5 and pH 8.0 with various substrates for selected species of basidiomycetes (A, B, C), ascomycete (D), commercially available lysine oxidase from T. viride (E) and V. ammodytes ammodytes venom (F). Following SDS-PAGE, LAO activity was detected by staining for H₂O₂ production using the OPD-HRP system and various LAO substrates: 1% tryptone (t), 0.1% CSM (c), 5 mM l-Lys (K), 5 mM l-Asn (N), 5 mM l-Phe (F), 5 mM l-Leu (L) in 0.1 M bis–Tris, pH 5.5, and 0.1 M Tris–HCl, pH 8.0 and control without substrate (ns) was performed at pH 5.5