A de novo designed N-terminal disulphide bridge stabilizes the Trichoderma reesei endo-1,4-β-xylanase II
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- 作者：Fenel ; Fred ; Leisola ; Matti ; Jä ; nis ; Janne ; Turunen ; Ossi
- 关键词：Endo-1 ; 4-β ; -xylanase ; Thermostability ; Thermophilicity ; N-terminus ; Disulphide bridge ; Trichoderma reesei
- 刊名：Journal of Biotechnology
- 出版年：2004
- 出版时间：March 4, 2004
- 年：2004
- 卷：108
- 期：2
- 页码：137-143
- 全文大小：123 K
文摘
We have successfully engineered a disulphide bridge into the N-terminal region of Trichoderma reesei endo-1,4-β-xylanase II (XYNII) by substituting Thr-2 and Thr-28 with cysteine. The T2C:T28C mutational changes increased the half-life in thermal inactivation of this mesophilic enzyme from 
40s to 20min at 65°C, and from less than 10s to 6min at 70°C. Therefore, the N-terminal disulphide bridge enables the use of XYNII at substantially higher temperatures than permitted by its native mesophilic counterpart. Altogether, thermostability increased by about 15°C. The kinetic properties of the mutant XYNII were maintained at the level of the wild type enzyme. Our findings demonstrated that a properly designed disulphide bridge, here within the N-terminal region of XYNII, can be very effective in resisting thermal inactivation.
40s to 20min at 65°C, and from less than 10s to 6min at 70°C. Therefore, the N-terminal disulphide bridge enables the use of XYNII at substantially higher temperatures than permitted by its native mesophilic counterpart. Altogether, thermostability increased by about 15°C. The kinetic properties of the mutant XYNII were maintained at the level of the wild type enzyme. Our findings demonstrated that a properly designed disulphide bridge, here within the N-terminal region of XYNII, can be very effective in resisting thermal inactivation.