Abstract Description

Microbial thermostable enzymes hold immense potential for industrial applications due to their resilience under harsh conditions. In this study, a thermostable lipase from an oil-contaminated soil bacterium, Bacillus velezensis, was cloned into E. coli BL21 for heterologous expression, purification, and characterisation. The truncated gene (500 bp) was amplified using gene-specific primers and cloned into a pET-28a(+) expression vector. The recombinant vector was successfully transformed into E. coli BL21 and the recombinant lipase was expressed using 0.4 mM IPTG as inducer at 30 °C after 7 hours of incubation. The enzyme was purified using Ni-NTA column chromatography with His-tag affinity and visualised as a 24 kDa protein on SDS-PAGE. The purified enzyme exhibited optimal activity at pH 9.0 and 80 °C. The recombinant lipase was most stable at 60 °C and pH 7, retaining 90 % and 95 % of its activity, respectively, after two hours. The properties of the enzyme suggest its potential for industrial applications, particularly in the detergent and biodiesel industries, and the degradation of oil-based pollutants.