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World J Microbiol Biotechnol. 2013 May;29(5):825-32. doi: 10.1007/s11274-012-1237-5. Epub 2012 Dec 21.

Expression and characterization of extreme alkaline, oxidation-resistant keratinase from Bacillus licheniformis in recombinant Bacillus subtilis WB600 expression system and its application in wool fiber processing.

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Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China.


A keratin-degrading bacterium of Bacillus licheniformis BBE11-1 was isolated and its ker gene encoding keratinase with native signal peptide was cloned and expressed in Bacillus subtilis WB600 under the strong P HpaII promoter of the pMA0911 vector. In the 3-L fermenter, the recombinant keratinase was secreted with 323 units/mL when non-induced after 24 h at 37 °C. And then, keratinase was concentrated and purified by hydrophobic interaction chromatography using HiTrap Phenyl-Sepharose Fast Flow. The recombinant keratinase had an optimal temperature and the pH at 40 °C and 10.5, respectively, and was stable at 10-50 °C and pH 7-11.5. We found this enzyme can retained 80 % activity after treated 5 h with 1 M H2O2, it was activated by Mg(2+), Co(2+) and could degraded broad substrates such as degraded feather, bovine serum albumin, casein, gelatin, the keratinase was considered to be a serine protease. Coordinate with Savinase, the keratinase could efficient prevent shrinkage and eliminate fibres of wool, which showed its potential in textile industries and detergent industries.

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