Purification, characterization, cDNA cloning, and expression of a xyloglucan endoglucanase from Geotrichum sp. M128

FEBS Letters

Yaoi, Katsuro¹  Mitsuishi, Yasushi¹ 

[1] Institute for Biological Resources and Functions, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba Central 6, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, Japan

关键词: Xyloglucanase;    Xyloglucan;    Endo-β-1;    4-glucanase;    XEG;    xyloglucan-specific endo-β-1;    4-glucanase;    OXG-RCBH;    oligoxyloglucan reducing end-specific cellobiohydrolase;    SDS–PAGE;    sodium dodecyl sulfate–polyacrylamide gel electrophoresis;    MALDI-TOF;    matrix-assisted laser desorption/ionization time-of-flight;    CMC;    carboxymethylcellulose;    RACE;    rapid amplification of cDNA ends;    CBM;    carbohydrate-binding module;   

DOI  :  10.1016/S0014-5793(04)00068-7

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

A novel xyloglucan-specific endo-β-1,4-glucanase (XEG), xyloglucanase, with a molecular mass of 80 kDa and a pI of 4.8, was isolated from the fungus Geotrichum sp. M128. It was found to be an endoglucanase active toward xyloglucan and not active toward carboxymethylcellulose, Avicel, or barley 1,3-1,4-β-glucan. Analysis of the precise substrate specificity using various xyloglucan oligosaccharide structures revealed that XEG has at least four subsites (−2 to +2) and specifically recognizes xylose branching at the +1 and +2 sites. The full-length cDNA encoding XEG was cloned and sequenced. It consists of a 2436-bp open reading frame encoding a 776-amino acid protein. From its deduced amino acid sequence, XEG can be classified as a family 74 glycosyl hydrolase. The cDNA encoding XEG was then expressed in Escherichia coli, and enzymatically active recombinant XEG was obtained.

【 授权许可】

Unknown