Construction of a Bacterial Assay for Estrogen Detection Based on an Estrogen-Sensitive Intein
Construction of a Bacterial Assay for Estrogen Detection Based on an Estrogen-Sensitive Intein
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基于雌激素敏感内含肽的雌激素检测细菌测定法的构建
DOI：
10.1128/aem.02336-10
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发表时间：
2011-02
影响因子：
4.4
通讯作者：
Liu, Jianhua
中科院分区：
文献类型：
--
作者：
Liang, Rubing;Zhou, Jing;Liu, Jianhua
通过在组成型表达的染色体lacZ基因的特异位点插入雌激素敏感内含肽（VMAER内含肽），构建了用于雌激素检测的大肠杆菌DIER菌株。该VMAER内含肽是通过将酿酒酵母VMA内含肽的核酸内切酶区域替换为人类雌激素受体α（hERα）的雌激素结合区域而产生的。当存在雌激素或类似物时，诱导VMAER内含肽的剪接以产生成熟的LacZ蛋白，其通过β-半乳糖苷酶比色测定来检测。利用该菌株对17-β-雌二醇、双酚A、芘、6-OH-芘、苯并[a]蒽、芘、孕酮和睾酮等8种典型化学物质进行了检测，整个检测过程在2 h内完成。它们的50%有效浓度（EC 50），相对雌激素活性，雌二醇当量因子进行了计算，并与酵母雌激素筛选（YES）系统检测到的结果是相当一致的。此外，对从上海某污水处理厂的废水和污泥中提取的人工麝香样品进行了雌激素活性检测，其结果与YES系统和气相色谱-质谱（GC-MS）的结果相当。总之，DIER生物测定法可以填补一个利基的有效，快速，高通量筛选雌激素化合物，并有潜力的远程，近实时监测环境雌激素。
ABSTRACT Escherichia coli strain DIER was constructed for estrogen detection by inserting an estrogen-sensitive intein (VMAER intein) into the specific site of the constitutively expressed chromosomal lacZ gene. This VMAER intein was generated by replacing the endonuclease region of the Saccharomyces cerevisiae VMA intein with the estrogen binding region of the human estrogen receptor α (hERα). When there were estrogens or analogs, the splicing of the VMAER intein was induced to produce the mature LacZ protein, which was detected through a β-galactosidase colorimetric assay. Eight typical chemicals (17-β-estradiol, bisphenol A, chrysene, 6-OH-chrysene, benz[a]anthracene, pyrene, progesterone, and testosterone) were detected using this DIER strain, and the whole detection procedure was accomplished in 2 h. Their 50% effective concentrations (EC50), relative estrogenic activities, and estradiol equivalency factors were calculated and were quite consistent with those detected with the yeast estrogen screening (YES) system. Furthermore, the estrogenic activities of the synthetic musk samples extracted from the wastewater and waste sludge of a sewage treatment plant of Shanghai (China) were detected, and their results were comparable to those obtained from the YES system and gas chromatography-mass spectrometry (GC-MS). In conclusion, the DIER bioassay could fill a niche for the efficient, rapid, high-throughput screening of estrogenic compounds and has potential for the remote, near-real-time monitoring of environmental estrogens.
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影响因子：
11.4
作者：
Layton, AC;Gregory, BW;Sayler, GS
通讯作者：
Sayler, GS
影响因子：
2.9
作者：
Brenzel, S;Kurpiers, T;Mootz, HD
通讯作者：
Mootz, HD
影响因子：
11.4
作者：
D. R. Raman;E. L. Williams;A. Layton;R. Burns;James P. Easter;Adam S. Daugherty;M. D. Mullen;G. Say
通讯作者：
D. R. Raman;E. L. Williams;A. Layton;R. Burns;James P. Easter;Adam S. Daugherty;M. D. Mullen;G. Say
影响因子：
11.4
作者：
T. Zacharewski
通讯作者：
T. Zacharewski
影响因子：
14.9
作者：
Perler, FB
通讯作者：
Perler, FB