Production of a novel cellulase by Bacillus amyloliquefaciens OKB3 isolated from soil: Purification and...

Microbial cellulases have become significant biocatalysts because of their complex composition and extensive industrial applications. This study aimed to isolate an efficient cellulase-producing strain, followed by molecular identification, enzyme purification, and characterization. Among 110 isolates, Bacillus amyloliquefaciens OKB3 was selected for its significant cellulase production, with optimal activity at pH 5.0 and 34 °C. The purification using ammonium sulfate and Sephadex G-100 chromatography resulted in specific activity of 2720.76 U/mg, 2.91 fold purification, and 29.44 % yield. The purified cellulase named CelB was a dimeric macromolecule of 123 kDa consisting of 67 and 54 kDa subunits. CelB was most active at 60 °C and pH 6, and it was stable at pH 5.5 to 6.0 and 0 °C to 4 °C. CelB was unaffected by metal cofactors and inhibited in the presence of divalent cations Cu²⁺, Hg²⁺, Cd²⁺, and Ag²⁺. The CelB has higher specificity of CMC compared to other substrates. The K_m, V_max, and K_cat values were 0.037 mM, 188.67 μmole/min, and 7430 S^-1 respectively. The unique attributes of CelB make it a very promising candidate for various biotechnological applications.