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Exploiting the Sensitivity of Nutrient Transporter Deletion Strains in Discovery of Natural Product Antimetabolites
ACS Infectious Diseases ( IF 4.0 ) Pub Date : 2017-11-07 00:00:00 , DOI: 10.1021/acsinfecdis.7b00149
Sebastian S. Gehrke 1 , Garima Kumar 1 , Nicole A. Yokubynas 1 , Jean-Philippe Côté 1 , Wenliang Wang 1 , Shawn French 1 , Craig R. MacNair 1 , Gerard D. Wright 1 , Eric D. Brown 1
Affiliation  

Actinomycete secondary metabolites are a renowned source of antibacterial chemical scaffolds. Herein, we present a target-specific approach that increases the detection of antimetabolites from natural sources by screening actinomycete-derived extracts against nutrient transporter deletion strains. On the basis of the growth rescue patterns of a collection of 22 Escherichia coli (E. coli) auxotrophic deletion strains representative of the major nutrient biosynthetic pathways, we demonstrate that antimetabolite detection from actinomycete-derived extracts prepared using traditional extraction platforms is masked by nutrient supplementation. In particular, we find poor sensitivity for the detection of antimetabolites targeting vitamin biosynthesis. To circumvent this and as a proof of principle, we exploit the differential activity of actinomycete extracts against E. coli ΔyigM, a biotin transporter deletion strain versus wildtype E. coli. We achieve more than a 100-fold increase in antimetabolite sensitivity using this method and demonstrate a successful bioassay-guided purification of the known biotin antimetabolite, amiclenomycin. Our findings provide a unique solution to uncover the full potential of naturally derived antibiotics.

中文翻译:

在天然产物抗代谢物的发现中开发营养转运蛋白缺失菌株的敏感性。

放线菌的次级代谢产物是抗菌化学支架的著名来源。在本文中,我们提出了一种针对特定目标的方法,该方法通过针对营养转运蛋白缺失菌株筛选放线菌来源的提取物,从而增加了从天然来源中抗代谢物的检测。基于22种大肠杆菌E. coli)集合的生长拯救模式)代表主要营养素生物合成途径的营养缺陷型菌株,我们证明了使用传统提取平台制备的放线菌来源提取物的抗代谢物检测被营养素补充掩盖了。特别是,我们发现检测针对维生素生物合成的抗代谢物的灵敏度较差。为避免这种情况并作为原理证明,我们利用放线菌提取物对大肠杆菌ΔyigM的差异活性,ΔyigM是一种生物素转运蛋白缺失菌株,相对于野生型大肠杆菌。使用这种方法,我们可以使抗代谢物的敏感性提高100倍以上,并证明了成功的生物测定指导下纯化了已知的生物素抗代谢物Amiclenomycin。我们的发现为发现天然抗生素的全部潜力提供了独特的解决方案。
更新日期:2017-11-07
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