| 中国鲎N-乙酰-β-D-氨基葡萄糖苷酶活性必需基团的研究 |
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| 引用本文： | 林建城,周文虎,林娟娟.中国鲎N-乙酰-β-D-氨基葡萄糖苷酶活性必需基团的研究[J].中国海洋大学学报(自然科学版),2020(1):57-64. |
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| 作者姓名： | 林建城 周文虎 林娟娟 |
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| 作者单位： | 莆田学院环境与生物工程学院福建省新型污染物生态毒理效应与控制重点实验室 |
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| 基金项目： | 福建省自然科学基金计划项目(2018J01467);福建省新型污染物生态毒理效应与控制重点实验室开放课题项目(PY16009)资助~~ |
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| 摘 要： | 为了探讨中国鲎N-乙酰-β-D-氨基葡萄糖苷酶(EC3.2.1.52,NAGase)活性的必需基团,采用了化学修饰法进行研究,分别以甲醛、乙酸酐、三硝基苯磺酸、对氯汞苯甲酸、苯甲基磺酰氟、二巯基苏糖醇、乙酰丙酮、溴代乙酸、碘代乙酸、N-溴代琥珀酰亚胺等10种修饰剂,在特定环境中对中国鲎NAGase进行化学修饰。结果表明:中国鲎NAGase中赖氨酸的ε-氨基、组氨酸的咪唑基、色氨酸的吲哚基和丝氨酸的羟基经修饰后,酶活力几近丧失,这些氨基酸基团为酶活性的必需基团,且可能处于酶的活性部位;半胱氨酸的巯基经修饰后不完全失活,说明半胱氨酸的巯基是NAGase催化活性所必需的,但可能不处于酶的活性部位;酶的二硫键被修饰后,酶活力明显下降,说明二硫键在维系NAGase的三维空间构象中起重要作用;而精氨酸的胍基经修饰后酶的活性基本不变,因此,精氨酸不是NAGase的必需基团。
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| 关 键 词： | 中国鲎 N-乙酰-β-D-氨基葡萄糖苷酶(NAGase) 化学修饰 必需基团 |
Study on the Essential Groups of the β-N-acetyl-D-glucosaminidase from the Viscera of Horseshoe Crab(Tachypleus tridentatus) |
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| LIN Jian-Cheng,ZHOU Wen-Hu,LIN Juan-Juan.Study on the Essential Groups of the β-N-acetyl-D-glucosaminidase from the Viscera of Horseshoe Crab(Tachypleus tridentatus)[J].Periodical of Ocean University of China,2020(1):57-64. |
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| Authors: | LIN Jian-Cheng ZHOU Wen-Hu LIN Juan-Juan |
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| Institution: | (College of Environmental and Biological Engineering,Fujian Provincial Key Laboratory of Ecology-toxicological Effects&Control for Emerging Contaminants,Putian University,Putian 351100,China) |
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| Abstract: | In order to identify the essential groups of β-N-acetyl-D-glucosaminidase(EC3.2.1.52) from the viscera of horseshoe crab(Tachypleus tridentatus), the chemical modification method had been used. The enzyme was modified respectively by ten chemical modification reagents, such as formaldehyde, acetic anhydride, trinitrobenzene sulfonic acid(TNBS), phenylmethanesulfonyl fluoride(PMSF), p-chloromercuribenzoate(pCMB), dithiothreitol(DTT), acetylaceton(AA), bromoacetic acid(BrAc), iodoacetic acid(IAc), N-bromosuccinimide(NBS) at certain condition. The results showed that the enzyme activity was almost lost when lysine’s ε-amido group, histidine’s imidazolyl group, tryptophan’s indolyl group and serine’s hydroxyl group were modified respectively, which means that these groups were essential for the enzyme catalytic activity, and probably situated in the active site of enzyme. The modification of cysteine’s sulfhydryl group led to inactivate incompletely, indicating sulfhydryl group was essential for the enzyme catalytic activity, but it might not be the composing groups of the enzyme active center. The enzyme activity was decreased significantly when the disulfide bond of enzyme were modified, which means that disulfide bond were essential for maintaining the enzyme’s conformation. The results also showed that the arginine’s guanidyl group was irrespective with the enzyme activity. |
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| Keywords: | Tachypleus tridentatus β-N-acetyl-D-glucosaminidase chemistry modification essential groups |
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