Engineering the thermostability of Trichoderma reesei endo-1,4-?-xylanase II by combination of disulphide bridges
- 20 July 2004
- journal article
- Published by Springer Nature in Extremophiles
- Vol. 8 (5) , 393-400
- https://doi.org/10.1007/s00792-004-0400-9
Abstract
Disulphide bridges were introduced in different combinations into the N-terminal region and the single α-helix of mesophilic Trichoderma reesei xylanase II (TRX II). We used earlier disulphide-bridge data and designed new disulphide bridges for the combination mutants. The most stable mutant contained two disulphide bridges (between positions 2 and 28 and between positions 110 and 154, respectively) and the mutations N11D, N38E, and Q162H. With a half-life of ~56 h at 65°C, the thermostability of this sevenfold mutant was ~5,000 times higher than that of TRX II, and the half-life was 25 min even at 75°C. The thermostability of this mutant was ~30 times higher than that of the corresponding mutant missing the bridge between positions 2 and 28. The extensive stabilization at two protein regions did not alter the kinetic properties of the sevenfold mutant from that of the wild-type TRX II. The combination of disulphide bridges enhanced significantly the pH-dependent stability in a wide pH range.Keywords
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