title: Characterization of endo-1,3-1,4-β-glucanases in GH family 12 from Magnaporthe oryzae.
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.
authors.affiliation: Iwate Biotechnology Research Center, Japan. ttakeda@ibrc.or.jp
authors.forename: Takumi
authors.initials: T
authors.lastname: Takeda
authors.forename: Machiko
authors.initials: M
authors.lastname: Takahashi
authors.forename: Tsugumi
authors.initials: T
authors.lastname: Nakanishi-Masuno
authors.forename: Yuki
authors.initials: Y
authors.lastname: Nakano
authors.forename: Hiromasa
authors.initials: H
authors.lastname: Saitoh
authors.forename: Akiko
authors.initials: A
authors.lastname: Hirabuchi
authors.forename: Shizuko
authors.initials: S
authors.lastname: Fujisawa
authors.forename: Ryohei
authors.initials: R
authors.lastname: Terauchi
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D058675:Brevibacillus / Q000201:enzymology / Q000235:genetics; D002473:Cell Wall / Q000378:metabolism; D002480:Cellulase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D004271:DNA, Fungal; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005800:Genes, Fungal; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006867:Hydrolases / Q000235:genetics; D020082:Magnaporthe / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism; D021382:Protein Sorting Signals; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D015398:Signal Transduction; D013379:Substrate Specificity; D047071:beta-Glucans / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004271:DNA, Fungal; D005656:Fungal Proteins; D011134:Polysaccharides; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D047071:beta-Glucans; D006867:Hydrolases; D006026:Glycoside Hydrolases; D002480:Cellulase; C028020:licheninase