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Applied and Environmental Microbiology, July 2002, p. 3496-3501, Vol. 68, No. 7
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.7.3496-3501.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Influence of the Transposition of the Thermostabilizing Domain of Clostridium thermocellum Xylanase (XynX) on Xylan Binding and Thermostabilization

Eun-Sun Shin,1 Mi-Jeong Yang,1 Kyung Hwa Jung,2 Eun-Ju Kwon,1 Jae Sung Jung,3 Seur Kee Park,4 Jungho Kim,1 Han Dae Yun,5 and Hoon Kim1*

Department of Agricultural Chemistry,1 Department of Biology,3 Department of Agricultural Biology, Sunchon National University, Sunchon 540-742,4 Amicogen, Inc., Chinsung, Chinju 660-852,2 Division of Applied Life Science, Gyeongsang National University, Chinju 660-701, Korea5

Received 12 February 2002/ Accepted 19 April 2002

A xylanase gene, xynX, of Clostridium thermocellum had one thermostabilizing domain (TSD) between the signal peptide sequence and the catalytic domain (CD). The TSD of a truncated xylanase gene, xynX'TSD-CD, was transpositioned from the N terminus to the C terminus of the CD by overlapping PCRs, and a modified product, xynX'CD-TSD, was constructed. XynX'TSD-CD had a higher optimum temperature (70°C versus 65°C) and was more thermostable (residual activity of 68% versus 46% after a 20-min preincubation at 70°C) than the one without the TSD, XynX'CD. However, the domain-transpositioned enzyme, XynX'CD-TSD, showed a lower optimum temperature (30°C) and thermostability (20%) than XynX'CD. Both XynX'TSD-CD and XynX'CD-TSD showed significantly higher binding capacity toward xylan than XynX'CD, and the domain transposition did not cause any change in the binding ability. XynX'TSD-CD and XynX'CD-TSD also showed considerable binding to lichenan but not to carboxymethyl cellulose and laminarin. XynX'TSD-CD and XynX'CD-TSD had higher activities for insoluble xylan than XynX'CD, while XynX'CD was more active against soluble xylan than XynX'TSD-CD and XynX'CD-TSD. These results indicate that the TSD of XynX has dual functions, xylan binding and thermostabilization, and the domain should also be classified as a xylan-binding domain (XBD). The binding capacity of the XBD was not affected by domain transpositioning within the gene.

* Corresponding author. Mailing address: Department of Agricultural Chemistry, Sunchon National University, Sunchon 540-742, Korea. Phone: 82-61-750-3294. Fax: 82-61-752-8011. E-mail: hoon{at}sunchon.ac.kr.

Applied and Environmental Microbiology, July 2002, p. 3496-3501, Vol. 68, No. 7
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.7.3496-3501.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.





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Copyright © 2002 by the American Society for Microbiology. All rights reserved.