Related Experiment Video
Updated: May 20, 2026
GENPLAT: an Automated Platform for Biomass Enzyme Discovery and Cocktail Optimization
Published on: October 24, 2011
Cloning and Heterologous Expression of a Novel Thermo-Alkalistable GH-10 Xylanase (rXyn-GM) Retrieved from Tapovan
Garima Chauhan1, Nivedita Bisht1, Praveen Gautam1
1Department of Biotechnology, School of Life Sciences, Babasaheb Bhimrao Ambedkar University, Lucknow, 226025 India.
Abstract:
A cellulase-free xylanase gene of 927 bp size (Xyn-GM) was isolated from the metagenomic library of the Tapovan Hot Spring in Uttarakhand, India. This gene encodes a 308-amino acid xylanase enzyme classified under the glycoside hydrolase family 10 (GH-10). The Xyn-GM gene was introduced into the pET28a ( +) vector and expressed in host cells of Escherichia coli BL21 (DE3). The recombinant xylanase (rXyn-GM), with a molecular weight ~ 32.5 kDa, was isolated through a one-step purification process using Ni2⁺-NTA affinity chromatography. The purified enzyme exhibited broad thermostability (50-100 °C) and pH stability (4.0-11.0), with optimal activity at 70 °C and pH 9.0. Its activity increased by 67% in the presence of 1 mM Mn2⁺. rXyn-GM retained ~ 65% activity after 2 h at 50 °C and 60 °C and ~ 75% activity at pH 9.0 after 3 h. It showed a preference for beechwood xylan, with kinetic parameters Km 20.9 mg/mL and Vmax 156.25 µmol/mg/min. Furthermore, rXyn-GM catalysed the production of xylo-oligosaccharides from beechwood xylan, suggesting its potential utility as prebiotics in the food and pharmaceutical industries.
Supplementary Information:
The online version contains supplementary material available at 10.1007/s12088-025-01480-1.
