標題: Cloning and Expression of Chitinase A from Serratia marcescens for Large-scale Preparation of N,N-Diacetyl Chitobiose
作者: Wu, Yue-Jin
Cheng, Chih-Yu
Li, Yaw-Kuen
應用化學系
Department of Applied Chemistry
關鍵字: Serratia marcescens;Chitinase A;Colloidal chitin;N,N-Diacetyl chitobiose
公開日期: 1-Aug-2009
摘要: The gene of Serratia marcescens chitinase A (chiA) was cloned by PCR. The complete gene was constructed into a pRSET vector and expressed in Escherichia coli. The recombinant enzyme was purified to > 90% homogeneity by hydrophobic interaction chromatography followed by ion-exchange separation. Measured with an electrospray-ionization mass spectrometer, the molecular mass of the protein was 58,607 Da, consistent with a theoretical calculation of the deduced protein without the signal peptide. The recombinant enzyme was characterized and tested for the preparation of chitobiose. In general, the recombinant Chtinase A exhibited an exo-type catalytic activity toward colloidal chitin and released both N-acetylglucosamine and N,N-diacetyl chitobiose as products. After extensive testing, we produced N,N-diacetyl chitobiose as the predominant product when the enzymatic reaction was performed in sodium acetate buffer at pH 5.5; under such conditions, an enzymatic process is established for the production of the disaccharide on a 100-g scale.
URI: http://hdl.handle.net/11536/6904
ISSN: 0009-4536
期刊: JOURNAL OF THE CHINESE CHEMICAL SOCIETY
Volume: 56
Issue: 4
起始頁: 688
結束頁: 695
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