[1]徐达,吴小卡,荆雅玮,等.布鲁菌半胱氨酸水解酶在甲硫氨酸循环中的催化活性研究[J].南京农业大学学报,2017,(4):697-702.[doi:10.7685/jnau.201608021]
 XU Da,WU Xiaoka,JING Yawei,et al.Catalytic activity of Brucella abortus SahH in activated methionine cycle[J].Journal of Nanjing Agricultural University,2017,(4):697-702.[doi:10.7685/jnau.201608021]
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布鲁菌半胱氨酸水解酶在甲硫氨酸循环中的催化活性研究()
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《南京农业大学学报》[ISSN:1000-2030/CN:32-1148/S]

卷:
期数:
2017年4期
页码:
697-702
栏目:
出版日期:
2017-07-07

文章信息/Info

Title:
Catalytic activity of Brucella abortus SahH in activated methionine cycle
作者:
徐达12 吴小卡1 荆雅玮1 左佳坤1 米荣升1 黄燕1 陈兆国1 王成明23 韩先干1
1. 中国农业科学院上海兽医研究所, 上海 200241;
2. 扬州大学兽医学院, 江苏 扬州 225009;
3. 奥本大学兽医学院, 美国 阿拉巴马州 36849-5519
Author(s):
XU Da12 WU Xiaoka1 JING Yawei1 ZUO Jiakun1 MI Rongsheng1 HUANG Yan1 CHEN Zhaoguo1 WANG Chengming23 HAN Xiangan1
1. Shanghai Veterinary Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China;
2. College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China;
3. Auburn University, Auburn AL 36849-5519, USA
关键词:
布鲁菌甲硫氨酸循环S-腺苷同型半胱氨酸水解酶催化活性信号分子AI-2
Keywords:
Brucella abortusactivated methionine cycleS-adenosylhomocysteine hydrolase(SahH)catalytic activityautoinducer-2
分类号:
Q343.1;S852.612
DOI:
10.7685/jnau.201608021
摘要:
[目的]细菌的sahH基因编码S-腺苷同型半胱氨酸水解酶(SahH),该酶参与细菌的甲硫氨酸循环(AMC),调控细菌的多种生理功能。[方法]通过构建重组表达质粒pET28a-Bru-sahH和pET28a-Pse-sahH,分别表达布鲁菌(Brucella abortus)S2308株和铜绿假单胞菌(Pseudomonas aeruginosa)PAO1株的SahH重组蛋白Bru-SahH和Pse-SahH。将纯化后的Bru-SahH、Pse-SahH以及我们前期表达纯化的禽致病性大肠杆菌(avian pathogenic Escherichia coli,APEC)的Pfs和LuxS蛋白,分别在体外催化S-腺苷同型半胱氨酸(SAH),通过对产物同型半胱氨酸(HCY)的浓度测定,评价不同重组蛋白的催化活性,并对催化底物时产生的自诱导分子2(AI-2)活性进行检测。[结果]Bru-SahH和Pse-SahH分别催化1 mmol·;L-1 SAH生成38和47 μmol·;L-1 HCY,而APEC的Pfs和LuxS蛋白能催化相同浓度的SAH产生401 μmol·;L-1 HCY。运用哈维弧菌BB170检测上述底物的AI-2活性,结果表明只有同时采用AEPC的Pfs和LuxS蛋白催化SAH,才能形成有活性的AI-2分子,而Bru-SahH和Pce-SahH均不能催化SAH形成活性AI-2分子。[结论]Bru-SahH能催化SAH生成HCY,为进一步研究sahH在布鲁菌感染过程中的作用提供依据。
Abstract:
[Objectives]The S-adenosylhomocysteine hydrolase(SahH)is encoded by sahH gene in Brucella abortus. SahH is a key enzyme in activated methionine cycle(AMC),which plays an important role in regulation of bacterial physiological functions. [Methods]In this study,the four recombinant proteins Bru-SahH(from B.abortus),Pse-SahH(from Pseudomonas aeruginosa),APEC-LuxS and APEC-Pfs(from avian pathogenic Escherichia coli)were purified respectively. The catalytic activity of the four recombinant proteins were evaluated by catalyzing substrate S-adenosylhomocysteine(SAH)to produce homocysteine(HCY)in vitro. [Results]The results showed that Bru-SahH or Pse-SahH could catalyze SAH(1 mmol·L-1)to produce 38 μmol·L-1 or 47 μmol·L-1 HCY,respectively,while APEC-LuxS and APEC-Pfs are 401 μmol·L-1. Furthermore,for confirming whether recombinant proteins could catalyze SAH to produce AI-2 or not,the detection of AI-2 bioassay by Vibrio harveyi strain BB170 was performed. The results demonstrated that APEC-LuxS and APEC-Pfs could catalyze SAH to produce AI-2,while Bru-SahH or Pse-SahH could not. [Conclusions]These findings will benefit for future studies on SahH regulation in Brucella abortus.

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备注/Memo

备注/Memo:
收稿日期:2016-08-19。
基金项目:国家自然科学基金项目(31572546,31370045);国家农产品质量安全风险评估计划项目(GJFP201700703)
作者简介:徐达,硕士研究生。
通信作者:韩先干,研究员,从事微生物致病机制、动物疫苗及动物疫病检测方法的研究,E-mail:hanxgan@163.com;王成明,教授,主要从事兽医分子诊断学、兽医微生物学和感染性肥胖的分子机制的研究,E-mail:wangche@auburn.edu。
更新日期/Last Update: 2017-07-07