title: Improved thermal performance of Thermomyces lanuginosus GH11 xylanase by engineering of an N-terminal disulfide bridge.
abstract: In order to increase the stability of thermophilic Thermomyces lanuginosus GH11 xylanase, TLX, a disulfide bridge Q1C-Q24C was introduced into the N-terminal region of the enzyme. The apparent temperature optimum shifted upwards at pH 6.5 by about 10°C to 75°C. The resistance to thermal inactivation also increased by about 10°C. The melting temperature measured by CD spectroscopy increased from 66 to 74°C. Therefore the N-terminal disulfide bridge increased both kinetic and thermodynamic stability almost equally. At pH 8 and 70°C, the disulfide bridge increased the enzyme half-life 20-fold in the presence of substrate. In contrast to the situation in acidic-neutral pH, the substrate decreased the thermostability of xylanases in alkaline pH. The upper limit for the performance of the disulfide bridge mutant at pH 9 was 75°C. This study showed that N-terminal disulfide bridges can stabilize even thermostable family GH11 xylanases.
authors.affiliation: Engineering Research Centre of Bioresources in Southern China, College of Life Science, South-central University for Nationalities, Wuhan 430074, China.
authors.forename: Yawei
authors.initials: Y
authors.lastname: Wang
authors.forename: Zheng
authors.initials: Z
authors.lastname: Fu
authors.forename: Huoqing
authors.initials: H
authors.lastname: Huang
authors.forename: Huashan
authors.initials: H
authors.lastname: Zhang
authors.forename: Bin
authors.initials: B
authors.lastname: Yao
authors.forename: Hairong
authors.initials: H
authors.lastname: Xiong
authors.forename: Ossi
authors.initials: O
authors.lastname: Turunen
mesh_terms: D000468:Alkalies; D001203:Ascomycota / Q000201:enzymology*; D002942:Circular Dichroism; D004220:Disulfides / Q000378:metabolism*; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry* / Q000378:metabolism*; D004789:Enzyme Activation; D004795:Enzyme Stability; D006207:Half-Life; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D009154:Mutation / Q000235:genetics; D015202:Protein Engineering / Q000379:methods*; D013696:Temperature; D013997:Time Factors; D044366:Transition Temperature
chemical_list: D000468:Alkalies; D004220:Disulfides; D043364:Endo-1,4-beta Xylanases