Resistance to a novel antichlamydial compound is mediated through mutations in Chlamydia trachomatis secY.
Resistance to a novel antichlamydial compound is mediated through mutations in Chlamydia trachomatis secY.
复制标题
对新型抗衣原体化合物的耐药性是通过沙眼衣原体 secY 的突变介导的。
DOI：
10.1128/aac.00356-12
复制
发表时间：
2012
影响因子：
4.9
通讯作者：
Rockey,DanielD
中科院分区：
文献类型：
--
作者：
Sandoz,KelsiM;Eriksen,StevenG;Jeffrey,BrendanM;Suchland,RobertJ;Putman,TimothyE;Hruby,DennisE;Jordan,Robert;Rockey,DanielD
探索了一种新的和定量的高通量筛选方法，作为鉴定抑制哺乳动物细胞中衣原体生长的新型化合物的工具。该测定基于细胞内衣原体对荧光标记物的积累。通过筛选42，000种化学上确定的化合物来证明其效用，以对抗衣原体GPIC。这一分析导致了40个主要命中化合物的鉴定。其中5个化合物对宿主细胞无毒性，对两种C.衣原体GPIC和沙眼衣原体。其中一种化合物（3-甲氧基苯基）-（4，4，7-三甲基-4，5-二氢-1H-[1，2]二硫杂环戊烯[3，4-C]喹啉-1-亚基）胺（MDQA）的抑制活性是衣原体特异性的，并被选择用于进一步研究。对MDQA的抗性筛选导致产生三个独立的抗性克隆。沙眼革兰氏阴性菌中参与Sec-dependent分泌的蛋白SecY的氨基酸变化与耐药表型相关。基于嗜热栖热菌SecY的已知结构，在每个抗性突变体中改变的氨基酸位于SecY晶体结构的预测中央通道中。这些实验模型的过程中，可用于发现抗衣原体，抗细胞内，或抗菌化合物，并导致鉴定的化合物，可能具有实用性的抗生素发现和进一步了解衣原体生物学。
A novel and quantitative high-throughput screening approach was explored as a tool for the identification of novel compounds that inhibit chlamydial growth in mammalian cells. The assay is based on accumulation of a fluorescent marker by intracellular chlamydiae. Its utility was demonstrated by screening 42,000 chemically defined compounds against Chlamydia caviae GPIC. This analysis led to the identification of 40 primary-hit compounds. Five of these compounds were nontoxic to host cells and had similar activities against both C. caviae GPIC and Chlamydia trachomatis. The inhibitory activity of one of the compounds, (3-methoxyphenyl)-(4,4,7-trimethyl-4,5-dihydro-1H-[1,2]dithiolo[3,4-C]quinolin-1-ylidene)amine (MDQA), was chlamydia specific and was selected for further study. Selection for resistance to MDQA led to the generation of three independent resistant clones of C. trachomatis. Amino acid changes in SecY, a protein involved in Sec-dependent secretion in Gram-negative bacteria, were associated with the resistance phenotype. The amino acids changed in each of the resistant mutants are located in the predicted central channel of a SecY crystal structure, based on the known structure of Thermus thermophilus SecY. These experiments model a process that can be used for the discovery of antichlamydial, anti-intracellular, or antibacterial compounds and has led to the identification of compounds that may have utility in both antibiotic discovery and furthering our understanding of chlamydial biology.