| Macromolecular Biosynthesis Assay for Evaluation of Influence of an Antimicrobial on the Synthesis of Macromolecules
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Author:
Date:
2013-06-20
[Abstract] One of the most compelling approaches in the discovery of novel antimicrobials is screening of natural sources. In our publication we report on the activity of a compound 8-hydroxyserrulat-14-en-19-oic acid (EN4), a diterpene isolated from the Australian plant Eremophila neglecta. We evaluate its applicability for treatment of implant-associated infections. A comprehensive analysis of the mechanism of action of EN4 against staphylococci revealed its membranolytic properties and a general inhibition of macromolecular biosynthesis, which was confirmed in a macromolecular biosynthesis assay and suggested a multitarget activity. The method used to investigate an influence of EN4 on the synthesis of peptidoglycan, RNA, DNA and proteins is based on precipitation of macromolecules with ...
[摘要] 在发现新的抗微生物剂中最引人注目的方法之一是筛选天然来源。在我们的出版物中，我们报告了一种化合物8-羟基胆固醇-14-烯-19-酸（EN4）（一种从澳大利亚植物中分离的二萜烯）的活性。我们评估其适用于植入物相关感染的治疗。对EN4对葡萄球菌的作用机制的综合分析揭示其膜分解性质和大分子生物合成的一般抑制，其在大分子生物合成测定中被证实并提出多靶点活性。用于研究EN4对肽聚糖，RNA，DNA和蛋白质的合成的影响的方法是基于大分子与三氯乙酸的沉淀。这些大分子由相应的[3 H] - 标记的前体合成。测量掺入和不掺入抗微生物剂的放射性，并反映测试化合物的作用模式。使用具有已知作用机制的抗生素作为对照。
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| Immunofluorescence Analysis of Yeast Protein
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Author:
Date:
2012-07-05
[Abstract] Many important regulatory proteins such as transcription factors are regulated through subcellular localization. Protein localization can be examined by fusing a GFP tag. However, GFP is relatively big in size, and potentially may affect correct protein localization. Several small tags have been developed, such as myc, HA or Flag. By using immunostain and fluorescence microscopy as described in this protocol, one can easily probe the regulation of a selected yeast protein with the application of the aforementioned small tags.
[摘要] 许多重要的调节蛋白如转录因子通过亚细胞定位进行调节。 蛋白质定位可以通过融合GFP标签来检查。 然而，GFP在尺寸上相对较大，并且潜在地可能影响正确的蛋白质定位。 已经开发了几个小标签，例如myc，HA或Flag。 通过使用如本方案中所述的免疫染色和荧光显微镜，可以通过应用上述小标签容易地探测所选择的酵母蛋白的调节。
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