title: Stability and conformational change of methoxypolyethylene glycol modification for native and unfolded trypsin.
abstract: The effect of succinimidyl carbonates activated methoxypolyethylene glycol (mPEG-SC) on the catalytic properties and conformation of native trypsin and dynamic high-pressure microfluidisation (DHPM) induced unfolded trypsin was studied. The thermal stability of unfolded trypsin was enhanced more significantly than that of native trypsin between 45 and 70°C. The autolysis analysis indicated that modified unfolded trypsin was markedly more resistant to autolysis compared to modified native trypsin between 40 and 180min. Upon mPEG-SC conjugation, the Km value of the enzyme decreased by about 2-fold, and the catalytic efficiency (Kcat/Km) increased by about 3-4-fold. Moreover, the increased thermal stability of unfolded trypsin might be due to the lower surface hydrophobicity and the higher hydrogen bond formation after mPEG-SC modification, which was reflected in the decrease of UV absorbance, the quenching and blue shift of fluorescence spectra, as well as the increase of β-sheet content.
authors.affiliation: State Key Laboratory of Food Science and Technology, Nanchang University, No. 235, Nanjing East Road, Nanchang, Jiangxi 330047, China. Electronic address: liuwei@ncu.edu.cn.
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.forename: Jun-Ping
authors.initials: JP
authors.lastname: Liu
authors.forename: Li-Qiang
authors.initials: LQ
authors.lastname: Zou
authors.forename: Zhao-Qin
authors.initials: ZQ
authors.lastname: Zhang
authors.forename: Cheng-Mei
authors.initials: CM
authors.lastname: Liu
authors.forename: Rui-Hong
authors.initials: RH
authors.lastname: Liang
authors.forename: Ming-Yong
authors.initials: MY
authors.lastname: Xie
authors.forename: Jie
authors.initials: J
authors.lastname: Wan
mesh_terms: D055162:Biocatalysis; D006358:Hot Temperature; D007700:Kinetics; D011092:Polyethylene Glycols / Q000737:chemistry*; D011487:Protein Conformation; D017510:Protein Folding; D055550:Protein Stability; D014357:Trypsin / Q000737:chemistry*
chemical_list: D011092:Polyethylene Glycols; C028210:monomethoxypolyethylene glycol; D014357:Trypsin
keywords: Conformational change; Methoxypolyethylene glycol (mPEG); Thermal stability; Trypsin