title: A novel alkaline surfactant-stable keratinase with superior feather-degrading potential based on library screening strategy.
abstract: A novel keratinase was mined and expressed in Escherichia coli BL21 (DE3) via function-driven screening with fosmid library. The catalytic properties of purified keratinase were investigated in detail following enzyme purification. The recombinant keratinase was purified to homogeneity with an estimated molecular weight of 26kDa using nickel affinity chromatography, of which the optimal reaction pH and temperature were 10.0 and 55°C, respectively. It could remain stable at pH 5.0-12.0 and 40-60°C. Metal ions such as Ca2+, Mn2+, Ag+, Na+, Mg2+, Li+, Sn2+(1mM) displayed positive influence on keratinase, and particularly, Ca2+exhibited remarkable improvement effect by 2.6 folds. It was strongly inhibited by PMSF as a protease inhibitor. On the contrary, it could be obviously activated by various surfactants, such as Tween 40 and Triton X-114. The recombinant keratinase showed high specificity towards casein, soluble keratin, BSA, and wool. The keratinase could efficiently degrade the feathers, which demonstrated its applicable potential for biodegradation of keratin wastes and regeneration of soluble protein.
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Chang
authors.initials: C
authors.lastname: Su
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Jin-Song
authors.initials: JS
authors.lastname: Gong
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Rong-Xian
authors.initials: RX
authors.lastname: Zhang
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Li-Yan
authors.initials: LY
authors.lastname: Tao
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Wen-Fang
authors.initials: WF
authors.lastname: Dou
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Dan-Dan
authors.initials: DD
authors.lastname: Zhang
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Heng
authors.initials: H
authors.lastname: Li
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Zhen-Ming
authors.initials: ZM
authors.lastname: Lu
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
authors.forename: Zheng-Hong
authors.initials: ZH
authors.lastname: Xu
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China. Electronic address: shijs@163.com.
authors.forename: Jin-Song
authors.initials: JS
authors.lastname: Shi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability / Q000187:drug effects; D005241:Feathers / Q000378:metabolism*; D015723:Gene Library; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000494:pharmacology; D008958:Models, Molecular; D010447:Peptide Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D013379:Substrate Specificity; D013501:Surface-Active Agents / Q000494:pharmacology*; D013696:Temperature
chemical_list: D004791:Enzyme Inhibitors; D008670:Metals; D013501:Surface-Active Agents; D010447:Peptide Hydrolases; C035855:keratinase
keywords: Characterization; Fosmid library; Keratinase