Modular Engineering to Enhance Keratinase Production for Biotransformation of Discarded Feathers

Biotransformation of wasted feathers via feather-degrading enzyme has gained immense popularity, low conversion efficiency hinders its scale application, and the main purpose of this study is to improve feather-degrading enzyme production in Bacillus licheniformis. Firstly, keratinase from Bacillus amyloliquefaciens K11 was attained with the best performance for feather hydrolysis, via screening several extracellular proteases from Bacillus; also, feather powder was proven as the most suitable substrate for determination of feather-degrading enzyme activity. Then, expression elements, including signal peptides and promoters, were optimized, and the combination of signal peptide SP_SacC with promoter Pdual3 owned the best performance, keratinase activity aggrandized by 6.21-fold. According to amino acid compositions of keratinase and feeding assays, Ala, Val, and Ser were proven as critical precursors, and strengthening these precursors' supplies via metabolic pathway optimization resulted in a 33.59% increase in the keratinase activity. Furthermore, keratinase activity reached 2210.66 U/mL, up to 56.74-fold from the original activity under the optimized fermentation condition in 3-L fermentor. Finally, the biotransformation process of discarded feathers by the fermented keratinase was optimized, and our results indicated that 90.94% of discarded feathers (16%, w/v) were decomposed in 12 h. Our results suggested that strengthening precursor amino acids' supplies was an efficient strategy for enhanced production of keratinase, and this research provided an efficient strain as well as the biotransformation process for discarded feather re-utilization.