title: Biochemical Characterization of An Arginine-Specific Alkaline Trypsin from Bacillus licheniformis.
abstract: In the present study, we isolated a trypsin-producing strain DMN6 from the leather waste and identified it as Bacillus licheniformis through a two-step screening strategy. The trypsin activity was increased up to 140 from 20 U/mL through culture optimization. The enzyme was purified to electrophoretic homogeneity with a molecular mass of 44 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the specific activity of purified enzyme is 350 U/mg with Nα-Benzoyl-L-arginine ethylester as the substrate. The optimum temperature and pH for the trypsin are 65 °C and pH 9.0, respectively. Also, the enzyme can be significantly activated by Ba(2+). This enzyme is relatively stable in alkaline environment and displays excellent activity at low temperatures. It could retain over 95% of enzyme activity after 180 min of incubation at 45 °C. The distinguished activity under low temperature and prominent stability enhance its catalytic potential. In the current work, the open reading frame was obtained with a length of 1371 nucleotides that encoded a protein of 456 amino acids. These data would warrant the B. licheniformis trypsin as a promising candidate for catalytic application in collagen preparation and leather bating through further protein engineering.
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. gjs713@jiangnan.edu.cn.
authors.forename: Jin-Song
authors.initials: JS
authors.lastname: Gong
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. qtdslw@163.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. zhangddlj@163.com.
authors.forename: Dan-Dan
authors.initials: DD
authors.lastname: Zhang
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. MinfengX@163.com.
authors.forename: Min-Feng
authors.initials: MF
authors.lastname: Xie
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. yb19920903@163.com.
authors.forename: Biao
authors.initials: B
authors.lastname: Yang
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. zrx_jiangnanedu@163.com.
authors.forename: Rong-Xian
authors.initials: RX
authors.lastname: Zhang
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. eternal83@163.com.
authors.forename: Heng
authors.initials: H
authors.lastname: Li
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. zmlu@jiangnan.edu.cn.
authors.forename: Zhen-Ming
authors.initials: ZM
authors.lastname: Lu
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. zhenghxu@jiangnan.edu.cn.
authors.forename: Zheng-Hong
authors.initials: ZH
authors.lastname: Xu
authors.affiliation: School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, China. shijs@163.com.
authors.forename: Jin-Song
authors.initials: JS
authors.lastname: Shi
mesh_terms: D000468:Alkalies / Q000378:metabolism*; D000595:Amino Acid Sequence; D001120:Arginine / Q000378:metabolism*; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D001483:Base Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability / Q000187:drug effects; D005285:Fermentation / Q000187:drug effects; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D007477:Ions; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D016415:Sequence Alignment; D013379:Substrate Specificity / Q000187:drug effects; D013696:Temperature; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000494:pharmacology
chemical_list: D000468:Alkalies; D007477:Ions; D008670:Metals; D012336:RNA, Ribosomal, 16S; D014361:Trypsin Inhibitors; D001120:Arginine; D014357:Trypsin
keywords: Bacillus licheniformis; biocatalysis; cloning; enzymatic properties; trypsin