Solid-state fermentation for protease production using agro-industrial residues by a novel thermotolerant...

The process of solid-state fermentation (SSF) offers a sustainable platform for enzyme production, particularly proteases with industrial relevance. In this study, protease synthesis was investigated using a thermotolerant Streptomyces sp. strain C8, isolated from arid soil of Tamanrasset, Algeria. The strain, Gram-positive with biverticillate spore chains, grew optimally at 45°C, pH 7, tolerated 5% NaCl, and exhibited 93.21% 16S rRNA similarity to Streptomyces griseorubens NBRC 12780 (GenBank accession PV834194), suggesting novelty. Under SSF, various agro-industrial wastes were tested, with spent brewing grains (SBGs) emerging as the most promising substrate. Initial optimization via the One-Factor-at-a-Time approach identified incubation time, temperature, pH, and moisture as critical parameters, yielding maximum activity (82 U/mL) under 30°C, pH 7, 4 days, and 75% humidity. Further optimization using response surface methodology of Box-Behnken design confirmed significant model performance (F = 8.47, P = 0.0003, R² = 0.9081), highlighting moisture, incubation time, and pH as key factors. The refined conditions (31°C, 55% moisture, 3 days, and pH 7) resulted in 128.70 U/mL activity. Overall, these findings demonstrate that low-cost agro-industrial byproducts, particularly SBGs, can serve as efficient, eco-friendly substrates for protease production using thermotolerant Streptomyces strains, offering promising applications in biotechnology.