Purification and characterization of a cold-active, organic solvent-, and surfactant-tolerant cellulase from <i>Pseudomonas fragi</i> HsL3-1 isolated from Hengshui Lake sediments

Cold-active cellulases attract significant attention for their potential in energy-efficient bioprocesses under low-temperature conditions. In this study, a psychrotolerant bacterial strain, Pseudomonas fragi HsL3-1, was isolated from Hengshui Lake sediments and found to produce a novel endoglucanase, EG-22SJ. The enzyme demonstrated optimal activity at pH 5.0 and 25°C, retaining over 80% and 60% of peak activity at 15°C and 5°C, respectively, and exhibited exceptional tolerance to 20% organic solvents (e.g. n-hexane enhanced activity by 29.8%) and 1% surfactants (e.g. Tween 80). Kinetic analysis revealed high substrate affinity for CMC-Na with a Km of 0.583 mg·ml-1 and Vmax of 401 μmol·l-1·min-1. Activity was significantly activated by Ca²⁺ and Mg²⁺ but inhibited by Cu²⁺ and Hg²⁺. Culture optimization via response surface methodology increased cellulase production to 8.71 U·ml-1 under conditions of 15.24 g·l-1 CMC-Na, 20.54°C, pH 6.85, and 1.95% inoculation, yielding a 1.24-fold improvement. These integrated properties position EG-22SJ as a robust biocatalyst for sustainable low-temperature applications such as biofuel production, food processing, and detergent formulation, highlighting the potential of nonextreme environments for enzyme discovery.