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Title 

Purification and characterization of Clostridium thermocellum xylanase from recombinant Escherichia coli

Authors 

Bon Joon KooHwa Gyun OhKi Haeng ChoChang Kun YangKyung Hwa JungDai Young Ryu

Publisher 

The Korean Society for Applied Microbiology

Issue Date 

1996

Citation 

Journal of Microbiology and Biotechnology, vol. 6, no. 6, pp. 414-419

Keywords 

clostridium thermocellumxylanase

Abstract 

The xylnX gene encoding a xylanase from Clostridium thermocellum ATCC27405 was cloned in the plasmid pJH27, an E. coli-Bacillus shuttle vector and the resultant recombinant plasmid, pJX18 was transformed into E. coli HB101. The overexpressed xylanase was found to be secreted into the periplasmic space of the recombinant E. coli cells. The crude enzyme was obtained by treating the E. coli cells with lysozyme, and purified by DEAE-Sepharose column chromatography. Molecular wieght of the xylanase was estimated to be 53 kDa by gel filtration. The pI value was determined to be pH 8.8. The N-terminal sequence of the enzyme protein was Asp-Asp-Asn-Asn-Ala-Asn-Leu-Val-Ser-Asn which was considered to be the sequence of that of the mature form protein. The Km value of the enzyme for oat spelt xylan was calculated to be 2.63 mg/ml and the Vmax value was 0.47 μmole/min. The xylanase had a pH optimum for its activity at pH 5.4 and a temperature optimum at 60°C. The enzyme hydrolyzed xylan into xylooligosaccharides which were composed mainly of xylobiose (40%) and xyloltriose (12%) after 5 hour reaction. This result indicates that the xylanase from C. thermocellum ATCC27405 is an endo-acting enzyme.

ISSN 

1017-7825

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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