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Applied Microbiology and Biotechnology

, Volume 88, Issue 5, pp 1113–1123 | Cite as

Characterization of endo-1,3–1,4-β-glucanases in GH family 12 from Magnaporthe oryzae

  • Takumi TakedaEmail author
  • Machiko Takahashi
  • Tsugumi Nakanishi-Masuno
  • Yuki Nakano
  • Hiromasa Saitoh
  • Akiko Hirabuchi
  • Shizuko Fujisawa
  • Ryohei Terauchi
Biotechnologically Relevant Enzymes and Proteins

Abstract

We have cloned three putative endoglucanase cDNAs, designated MoCel12A, MoCel12B, and MoCel12C, from Magnaporthe oryzae. The deduced peptide sequences of both MoCel12A and MoCel12B contain secretion signal peptides and a catalytic core domain that classify them into GH subfamily 12-1. In contrast, the deduced peptide sequence of MoCel12C consists of a signal peptide, a catalytic core domain, and a fungal-type carbohydrate binding module belonging to GH subfamily 12-2. Although most GH family 12 endoglucanases hydrolyze β-1,4-glucans such as carboxymethylcellulose or phosphoric acid-swollen cellulose, MoCel12A that was prepared by overexpression in M. oryzae and Brevibacillus choshinensis hydrolyzed specifically 1,3–1,4-β-glucans, such as barley β-glucan and lichenan. The specific activity of MoCel12A overexpressed in M. oryzae was about 20 times higher than that prepared from B. choshinensis. Furthermore, MoCel12B prepared by overexpression in B. choshinensis also revealed preferential hydrolysis of endo-1,3–1,4-β-glucans with limited hydrolysis on carboxymethylcellulose. In comparison with MoCel12A, the activity of MoCel12B was more stable under alkaline conditions. Levels of mRNA encoding MoCel12A were constitutively high during infection and spore formation. The overexpression and disruption of the MoCel12A gene did not affect germination, appressorium formation, or invasion rate; however, M. oryzae overexpressing MoCel12A produced larger numbers of spores than the wild type or a mutant in which the MoCel12A gene was disrupted. These results suggest that MoCel12A functions in part to hydrolyze 1,3–1,4-β-glucan during infection and spore formation.

Keywords

Endo-1,3–1,4-β-glucanase β-glucan hydrolysis GH family 12 Magnaporthe oryzae Overexpression Spore formation 

Notes

Acknowledegments

This work was supported in part by the New Energy and Industrial Technology Development Organization of Japan (NEDO).

Supplementary material

253_2010_2781_MOESM1_ESM.pdf (2.3 mb)
ESM (PDF 2337 kb)

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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Takumi Takeda
    • 1
    Email author
  • Machiko Takahashi
    • 1
  • Tsugumi Nakanishi-Masuno
    • 1
  • Yuki Nakano
    • 1
  • Hiromasa Saitoh
    • 1
  • Akiko Hirabuchi
    • 1
  • Shizuko Fujisawa
    • 1
  • Ryohei Terauchi
    • 1
  1. 1.Iwate Biotechnology Research CenterKitakamiJapan

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