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- Title
Characterization of antifungal activity of the GH-46 subclass III chitosanase from Bacillus circulans MH-K1.
- Authors
Tomita, Masayo; Kikuchi, Ayami; Kobayashi, Mariko; Yamaguchi, Masashi; Ifuku, Shinsuke; Yamashoji, Shiro; Ando, Akikazu; Saito, Akihiro
- Abstract
We characterized the antifungal activity of the Bacillus circulans subclass III MH-K1 chitosanase (MH-K1 chitosanase), which is one of the most intensively studied glycoside hydrolases (GHs) that belong to GH family 46. MH-K1 chitosanase inhibited the growth of zygomycetes fungi, Rhizopus and Mucor, even at 10 pmol (0.3 μg)/ml culture probably via its fungistatic effect. The amino acid substitution E37Q abolished the antifungal activity of MH-K1 chitosanase, but retained binding to chitotriose. The E37Q mutant was fused with green fluorescent protein (GFP) at its N-terminus and proved to act as a chitosan probe in combination with wheat-germ agglutinin (WGA), which is a chitin-specific binding lectin. The GFP-fused MH-K1 chitosanase mutant E37Q (GFP-E37Q) bound clearly to the hyphae of the Rhizopus and Mucor strains, indicating the presence of chitosan. In contrast, Cy5-labelled WGA (Cy5-WGA), but not GFP-E37Q, stained the hyphae of non-zygomycetes species, i.e. Fusarium oxysporum, Penicillium expansum, and Aspergillus awamori. When the mycelia of Rhizopus oryzae were treated with wild type MH-K1 chitosanase, they could not bind to GFP-E37Q but were stained instead by Cy5-WGA. We conclude that chitin is covered by chitosan in the cell walls of R. oryzae.
- Subjects
BACILLUS circulans; GLYCOSIDASES; GENETIC mutation; MUCOR; FUSARIUM oxysporum; APPLE blue mold
- Publication
Antonie van Leeuwenhoek, 2013, Vol 104, Issue 5, p737
- ISSN
0003-6072
- Publication type
Article
- DOI
10.1007/s10482-013-9982-5