High-Level Extracellular Expression of Collagenase ColH in Bacillus subtilis for Adipose-Derived Cells Extraction

Collagenase has a wide range of applications in the medicine, cosmetic, and food industries. Inefficient expression of collagenase impedes its industrial production and commercial applications. In this study, a secretory expression system for collagenase ColH from Clostridium histolyticum was constructed in Bacillus subtilis. Signal peptide optimization effectively solved the secretion problem of large collagenase with a molecular weight of about 116 kDa, doubling the extracellular enzyme activity. Then, promoter optimization further improved the enzyme activity to 264 U/mL. By the co-optimization of the nitrogen sources and carbon sources, and employing a fed-batch fermentation strategy, the enzyme activity could reach 669 U/mL, which is, currently, the highest level reported in the industry. The recombinant collagenase ColH was purified through a purification process suitable for industrial production with a specific activity of 565.25 U/mg. Based on the purified collagenase, cells were successfully prepared from adipose tissue, indicating its potential use in cell therapy. This study provides a promising candidate for the industrial production of collagenase and highlights its potential application to extract cells from tissues.