Inhibitory Effect of Dimethylsulfoxide on N-Acetyl-β-D-Glucosaminidase from Chinese Horseshoe Crab Tachypleus tridentatus
LIN Jiancheng, WU Jianhong, LIN Juanjuan
Key Laboratory of Ecological Environment and Information Atlas (Putian University), Fujian Provincial Key Laboratory of Ecology-Toxicological Effects & Control for Emerging Contaminants, College of Environmental and Biological Engineering, Putian University, Putian 351100, China
Abstract：The periodic molting of arthropods is closely related to N-acetyl-β-D-glucosaminidase (EC3.2.1.52, NAGase). NAGase was extracted from the viscera of Tachypleus tridentatus. The effect of dimethyl sulfoxide (DMSO) on NAGase was studied by using the kinetics method of enzymatic reaction. The results showed that DMSO had strong inhibitory effect on the NAGase, the enzyme activity decreased by 38.0% when its concentration reached 0.1 mol/L. The inhibitory effect of DMSO on NAGase was reversible. The kinetics showed that the inhibitory effect of DMSO solution on NAGase was the competitive and non-competitive mixed type. The inhibition constants of DMSO on free enzyme (KI) and on enzyme-substrate complex(KIS) were determined to be 0.200 mol/L and 1.063 mol/L, respectively. The value of KI was less thanthat of KIS, indicating a marked protective effect of the substrate on the inactivation of the enzyme. As the concentration of substrate increased, the inactivation of enzyme in DMSO turned to decrease. Meanwhile, UV absorption spectra and fluorescence emission spectra of NAGase were measured. The UV absorption of NAGase treated by DMSO was enhanced with increasing DMSO concentration, but the fluorescence emission peak intensity of the enzyme gradually decreased, their UV absorption peak and fluorescence emission peak of NAGase did not shift. The fluorescence emission of enzyme was almost quenched when the concentration of DMSO reached 0.65 mol/L. It showed that changes of enzyme conformation in the DMSO solvent led to inactivation of the NAGase. The above results confirmed that DMSO might further affect its molting by inhibiting the activity of NAGase from Tachypleus tridentatus.
林建城, 吴建洪, 林娟娟. 二甲亚砜对中国鲎NAGase的抑制作用[J]. 水产科学, 2023, 42(4): 719-725.
LIN Jiancheng, WU Jianhong, LIN Juanjuan. Inhibitory Effect of Dimethylsulfoxide on N-Acetyl-β-D-Glucosaminidase from Chinese Horseshoe Crab Tachypleus tridentatus. 水产科学, 2023, 42(4): 719-725.
[1]刘树深,刘芳,刘海玲.20种水溶性有机溶剂对发光菌的毒性效应[J].中国环境科学,2007,27(3):371-376. [2]YANG G P, WANG X, ZHANG H H, et al. Temporal and spatial variations of dimethylsulfoxide in the Bohai Sea and the Yellow Sea[J].Journal of Sea Research,2014,90:33-43. [3]王鑫,张洪海,杨桂朋.东海和黄海冬季海水中二甲亚砜的分布及影响因素[J].环境科学研究,2014,27(10):1119-1125. [4]王英红,徐虹,孔庆鑫.低浓度二甲基亚砜对斑马鱼胚胎发育和成鱼生殖的影响[J].中国卫生检验杂志,2020,30(6):662-665. [5]BOWNIK A. Effects of ectoine on behavioral, physiological and biochemical parameters of Daphnia magna exposed to dimethyl sulfoxide[J].Science of the Total Environment,2019,683:193-201. [6]龚敏,陈清西,林建城,等.太平洋白对虾(Penaeus vannamei)β-N-乙酰-D-氨基葡萄糖苷酶在二甲亚砜溶液中的失活动力学[J].中国生物化学与分子生物学报,2005,21(5):581-585. [7]张继平,陈清西.锯缘青蟹NAGase在二甲亚砜溶液中的失活动力学[J].台湾海峡,2006,25(1):19-24. [8]谢进金,陈丽娟.二甲亚砜对黄粉虫NAGase活力的影响[J].泉州师范学院学报,2006,24(4):96-100. [9]ZOU E, FINGERMAN M. Chitobiase activity in the epidermis and hepatopancreas of the fiddler crab Uca pugilator during the molting cycle[J].Marine Biology,1999,133(1):97-101. [10]黄乾生,谢晓兰,石艳,等.不同生长期凡纳滨对虾肝胰腺NAGase的基本性质变化[J].厦门大学学报(自然科学版),2006,45(6):847-850. [11]林建城,王丽英,林娟娟.中国鲎几丁质酶的酶学性质研究[J].水产科学,2019,38(5):702-709. [12]林建城,许恒棋,罗新明.中国鲎内脏N-乙酰-β-D-氨基葡萄糖苷酶的分离纯化及其酶学性质[J].中国海洋大学学报(自然科学版),2017,47(9):62-69. [13]ZHANG W N, CHEN Q X, LIN X Y, et al. Irreversible inhibitory kinetics of mercuric ion on N-acetyl-β-D-glucosaminidase from Nile tilapia (Oreochromis niloticus)[J].Aquatic Toxicology,2014,154:163-167. [14]陈清西.酶学及其研究技术[M].2版.厦门:厦门大学出版社,2015:102,120,250-257. [15]梁娟然,戴菲辉,苏思莹,等.纤维素酶在二甲基亚砜微扰条件下的动力学行为和光谱学变化[J].生物加工过程,2011,9(2):29-32. [16]SANA B, GHOSH D, SAHA M, et al. Purification and characterization of an extremely dimethylsulfoxide tolerant esterase from a salt-tolerant Bacillus species isolated from the marine environment of the Sundarbans[J].Process Biochemistry,2007,42(12):1571-1578. [17]ALSAFADI D, PARADISI F. Effect of organic solvents on the activity and stability of halophilic alcohol dehydrogenase (ADH2) from Haloferax volcanii[J].Extremophiles,2013,17(1):115-122. [18]FAULDS C B, PÉREZ-BOADA M, MARTíNEZ Á T. Influence of organic co-solvents on the activity and substrate specificity of feruloyl esterases[J].Bioresource Technology,2011,102(8):4962-4967. [19]刘永萍,杨平,王岚,等.二甲基亚砜对CAOV3细胞端粒酶活性的可逆性抑制作用[J].中国现代医学杂志,2006,16(21):3241-3244. [20]CHEN Q X, LIU X D, HUANG H. Inactivation kinetics of mushroom tyrosinase in the dimethyl sulfoxide solution[J].Biochemistry (Moscow),2003,68(6):644-649. [21]MANGIAGALLI M, CARVALHO H, NATALELLO A, et al. Diverse effects of aqueous polar co-solvents on Candida antarctica lipase B[J].International Journal of Biological Macromolecules,2020,150:930-940. [22]MUKHERJEE A, SARKAR S, GUPTA S, et al. DMSO strengthens chitin deacetylase-chitin interac-tion: Physicochemical, kinetic, structural and catalytic insights[J].Carbohydrate Polymers,2019,223:115032. [23]黎春怡,黄卓烈,李利佳,等.二甲基亚砜和四氢呋喃对胃蛋白酶催化动力学和分子光谱的影响[J].高等学校化学学报,2012,33(5):988-995.
2023, Vol. 42 
