1887

Abstract

A chitosanase was purified from the intercellular fluid extract of chemically stressed barley () leaves. Purification was achieved by preparative PAGE involving separation under native conditions at pH 4.3 (Reisfeld system) followed by denaturing PAGE in the presence of SDS. One basic chitosanase with a molecular mass of 19 kDa could hydrolyse chitosan and glycol chitosan of crustacean origin in addition to fungal chitosan isolated after treatment of cells with sodium hydroxide followed by extraction in acetic acid. This enzyme did not exhibit activity towards -1,4-polymers such as chitin, cellulose or peptidoglycan (from ) after SDS-PAGE. The chitosanase was conjugated to colloidal gold at pH 9·5 and applied to fungal tissue sections or to isolated glycol chitosan, chitosan, cellulose or chitin. The chitosanase-gold complex reacted intensely with glycol chitosan and chitosan but did not bind to chitin or cellulose. The enzyme-gold complex also labelled chitosan isolated from . However, it did not react with cells in tissue sections. Labelling with the chitosanase-gold complex was easily detected over cell walls of f. sp. spores and hyphae as well as over cell walls of and . No binding was observed with .

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1991-08-01
2024-12-02
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