Purification, characterization and physiological significance of a chitinase from the pilei of Coprinopsis cinerea fruiting bodies

被引:29
作者
Zhou, Yajun [1 ]
Kang, Liqin [1 ]
Niu, Xin [1 ]
Wang, Jun [1 ]
Liu, Zhonghua [1 ]
Yuan, Sheng [1 ]
机构
[1] Nanjing Normal Univ, Coll Life Sci, Jiangsu Engn & Technol Res Ctr Industrializat Mic, Jiangsu Key Lab Microbes & Microbial Funct Genom, Nanjing 210023, Jiangsu, Peoples R China
基金
中国国家自然科学基金;
关键词
chitinase; cell wall; pileus; autolysis; fruiting body; CELL-WALL; TRICHODERMA-HARZIANUM; SYNERGISTIC ACTIVITY; AUTOLYSIS; CHITOBIOSIDASE; MUSHROOM; ENZYMES; CLONING;
D O I
10.1093/femsle/fnw120
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
We purified a chitinase from pilei extractions of Coprinopsis cinerea fruiting bodies by ammonium sulfate precipitation and CM Sepharose cation exchange chromatography. MALDI-TOF/TOF MS analysis characterized this purified chitinase as a putative class V chitinase, ChiB1. ChiB1 hydrolyzed colloidal chitin and chitosan, whereas it did not hydrolyze chitin powder. ChiB1 cleaved only pNP-(GlcNAc)(2), rather than pNP-GlcNAc or pNP-(Glc-NAc)(3), to release nitrophenol. ChiB1 preferably and progressively released (GlcNAc)(2) from (GlcNAc) 6 and digested (GlcNAc)6 to two molecules of (GlcNAc)3 in a small proportion, but did not split (GlcNAc)(2), so it is an exochitinase. ChiB1 has an optimum temperature range of 35 degrees C to 40 degrees C and an optimum pH of 5.0. ChiB1 exhibited Km and Vmax values of 2.63 mg ml(-1) and 2.31 mu mol min(-1) mg protein(-1) for colloidal chitin, respectively. The ChiB1 gene, along with another putative endochitinase (class III chitinase gene), was expressed dominantly among eight predicted chitinase genes in the genome, and its expression level increased with the maturation of fruiting bodies. ChiB1 incubation released a large amount of soluble beta-glucan fractions from alkali-insoluble cell wall fractions of C. cinerea fruiting bodies, thereby it may promote the degradation of cell walls in synergy with the beta-1,3-glucanases during pileus autolysis.
引用
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页数:7
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