Isolation, genome mining, and statistical optimization of keratinase production by Bacillus paralicheniformis SuvM...

In this study, a bacterium was isolated from a Himalayan hot spring of Gangnani, Uttarakhand, India, capable of producing keratinase. The whole-genome sequencing and phylogenetic analysis identified it as Bacillus paralicheniformis SuvM (JBQZRG000000000). Genome annotation confirmed the presence of a putative keratinase gene harboring a signal peptide, an inhibitor I-9 region, and a peptidase S8 catalytic domain, characteristic of subtilisin-like serine proteases with molecular weight of 38.867 kDa. The crude enzyme extract exhibited maximum keratinolytic activity of 425.33 U/mg protein (∼70 U/mL). To enhance enzyme production, Response Surface Methodology (RSM) was employed with process variables: temperature, pH, incubation time, and agitation speed. A statistically significant quadratic model was obtained with a probability of less than 0.0001 and with a high coefficient of determination (R² = 0.98), confirming its robustness for predictive optimization. Interaction analysis revealed synergistic effects of temperature and pH, while optimum conditions were determined as 50 °C, pH 7.5, 120 RPM, and 48 h. Keratinase activity increased markedly under these conditions compared to unoptimized controls. In general, Bacillus paralicheniformis SuvM represents a promising source of thermophilic keratinase. The optimized process harnesses its potential for applications in feather degradation, waste valorization, and sustainable bioprocesses, therefore justifying further exploration of hot spring isolates as reservoirs of industrially important enzymes.