Tomas Richardson-Sanchez, Thomas J Telfer, Cho Z Soe, Kate P Nolan, Michael P Gotsbacher, Rachel Codd
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引用次数: 0
摘要
嗜苷酸盐是细菌和真菌的低分子量有机次级代谢产物,可与铁(III)形成高亲和力络合物。这些铁(III)-苷元复合物是苷元介导的铁(III)吸收机制的一部分,是微生物为获得足够的铁以促进生长而使用的最广泛的策略。微生物对有限铁元素的竞争是通过生物合成基因簇来解决的,这些基因簇可编码具有不同分子支架和铁结合基团的嗜苷酸的生物合成。某些类别的嗜苷酸的生物合成途径已广为人知,这为利用前体定向生物合成(PDB)进一步扩大结构和性质的多样性提供了机会。PDB 包括在培养基中添加非原生底物,以便在代谢物组装过程中与原生底物竞争。本章介绍了为生产一系列不同的原型羟肟酸苷酸盐去铁胺 B 类似物而进行 PDB 实验的背景信息和技术细节,包括半纯化培养上清液的过程,以及使用液相色谱-串联质谱法对类似物和组构异构体进行下游分析的过程。
The production of siderophore analogues using precursor-directed biosynthesis.
Siderophores are low-molecular-weight organic bacterial and fungal secondary metabolites that form high affinity complexes with Fe(III). These Fe(III)-siderophore complexes are part of the siderophore-mediated Fe(III) uptake mechanism, which is the most widespread strategy used by microbes to access sufficient iron for growth. Microbial competition for limited iron is met by biosynthetic gene clusters that encode for the biosynthesis of siderophores with variable molecular scaffolds and iron binding motifs. Some classes of siderophores have well understood biosynthetic pathways, which opens opportunities to further expand structural and property diversity using precursor-directed biosynthesis (PDB). PDB involves augmenting culture medium with non-native substrates to compete against native substrates during metabolite assembly. This chapter provides background information and technical details of conducting a PDB experiment towards producing a range of different analogues of the archetypal hydroxamic acid siderophore desferrioxamine B. This includes processes to semi-purify the culture supernatant and the use of liquid chromatography-tandem mass spectrometry for downstream analysis of analogues and groups of constitutional isomers.
期刊介绍:
The critically acclaimed laboratory standard for almost 50 years, Methods in Enzymology is one of the most highly respected publications in the field of biochemistry. Each volume is eagerly awaited, frequently consulted, and praised by researchers and reviewers alike. Now with over 500 volumes the series contains much material still relevant today and is truly an essential publication for researchers in all fields of life sciences, including microbiology, biochemistry, cancer research and genetics-just to name a few. Five of the 2013 Nobel Laureates have edited or contributed to volumes of MIE.