Engineering a high-performance, metagenomic-derived novel xylanase with improved soluble protein yield and thermostability

• A novel metagenomic-derived xylanase gene xyl7 was isolated and expressed.
• The enzyme XYL7 showed higher specific activity and broad pH activity range.
• A truncated XYL7-Tr3 with tenfold increase in soluble protein yield was acquired.
• Mutants showed 250-fold and 10 °C increase in half-life and optimal temperature.
• The reducing sugar produced from xylan by mutant XYL7-TC was increased fourfold.

The novel termite gut metagenomic-derived GH11 xylanase gene xyl7 was expressed in Escherichia coli BL21, and the purified XYL7 enzyme exhibited high specific activity (6340 U/mg) and broad pH active range of 5.5–10.0. Directed evolution was employed to enhance the thermostability of XYL7; two mutants (XYL7-TC and XYL7-TS) showed a 250-fold increase in half-life at 55 °C, with a 10 °C increase in optimal temperature compared to that of wild-type XYL7. A truncated enzyme (XYL7-Tr3) acquired by protein engineering showed similar catalytic properties as the wild-type, with a tenfold increase in soluble protein yield by the mutant. The reducing sugar produced by XYL7-TC was about fourfold greater than that produced by their parents when incubated with xylan at 60 °C for 4 h. The engineered novel xylanase exhibited superior enzymatic performance and showed promise as an excellent candidate for industrial application due to its high specific activity, stability and soluble protein yield.