Expression of a Trichoderma reesei beta-xylanase gene in Escherichia coli and activity of the enzyme on fiber-bound substrates

Protein Expression and Purification
He JunChen Daiwen

Abstract

A recombinant gene Xyn2 (570bp) encoding the main Trichoderma reesei Rut C-30 endo-beta-1,4-xylanase was successfully cloned and expressed in Escherichia coli BL21 under the control of strong bacteriophage T7 transcription and translation signals. The molecular weight of the recombinant protein was estimated by SDS-PAGE to be 24 kDa. The expressed protein was purified by Ni(2+)-NTA affinity chromatography and enzyme activity assay verified the protein as a xylanase. Like with the T. reesei Xyn2, the optimal activity of the recombinant Xyn2 was at 50 degrees C. However, the recombinant xylanase had an improved thermostability and more than 70% of its activity remained after 30 min incubation at 60 degrees C. In addition, the recombinant xylanase was active over the range of pH 3.5-7.5 with maximum activity at pH 5.0. The enzyme was highly specific towards xylans but exhibited very low activities towards cellulosic substrates. Using Birchwood xylan, the determined apparent K(m) and k(cat) values were 0.11 mg/ml and 106 s(-1), respectively. These properties should make the enzyme an attractive candidate for various industrial applications.

References

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Citations

Feb 9, 2011·Applied Biochemistry and Biotechnology·Peng ZhouZhengqiang Jiang
Jun 22, 2013·Journal of Basic Microbiology·Wei FangAnshan Shan
May 6, 2015·International Journal of Biological Macromolecules·Fatma ElgharbiSamir Bejar
Dec 12, 2012·Bioengineered·Hui LinYuhua Zhao
Dec 23, 2014·International Journal of Biological Macromolecules·Fatma ElgharbiAïda Hmida-Sayari
Jul 2, 2009·Applied Microbiology and Biotechnology·Sibtain AhmedAmer Jamil
Mar 6, 2017·Microbial Cell Factories·Laia GifreElena Garcia-Fruitós
Mar 23, 2013·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Qun WanPaul Langan

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