Biochemical and functional characterization of a novel thermoacidophilic, heat and halo-ionic liquids tolerant endo-β-1,4-glucanase from saline-alkaline lake soil microbial metagenomic DNA

The lignocellulosic biomass usually need pre-treatment with acid at high temperature or solved in ionic liquid (IL) before cellulases hydrolysis. Thus, thermoacidophilic, thermostable and IL-tolerant cellulases are greatly desired, but rarely reported. In this study, a novel endo-β-1,4-glucanase gene nmGH45 was directly cloned from saline-alkaline lake soil microbial metagenomic DNA and expressed in Pichia pastoris. Recombinant NMgh45 was active and stable at pH 3.0-9.0, with maximum activity at pH 4.5 and >80% residual activities at pH 3.0-11.0. It exhibited maximal activity at 60-70 °C and remaining >90% and 68% residual activities at 80 °C for 2 h and 90 °C for 1 h. Besides, NMgh45 retained >88% relative activities in 4 M NaCl, and was stable for 24 h. Significantly, it was highly IL-tolerant, remaining 92.8% and 43.8% residual activities in 10% and 20% 1-butyl-3-methylimidazolium chloride. Compared with other non-halo-tolerant counterparts, NMgh45 was enriched with acidic amino acids on the protein surface, and the conformation of NMgh45 was stable at high salinity condition based on molecular dynamic simulation. Finally, the productivity of recombinant NMgh45 was optimized at multi-levels. The thermoacidphilic and acid-alkaline-halo-tolerant properties make NMgh45 a promising enzyme for basic research and industrial applications.