Affinity purification method for the identification of nonribosomal peptide biosynthetic enzymes using a synthetic probe for adenylation domains
Affinity purification method for the identification of nonribosomal peptide biosynthetic enzymes using a synthetic probe for adenylation domains
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使用腺苷酸化结构域合成探针鉴定非核糖体肽生物合成酶的亲和纯化方法
DOI：
10.1007/978-1-4939-3375-4_4
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发表时间：
2016
期刊：
影响因子：
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通讯作者：
Fumihiro Ishikawa and Hideaki Kakeya
中科院分区：
文献类型：
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作者：
杉山 康憲;亀下 勇;坂本 修士;Fumihiro Ishikawa and Hideaki Kakeya
基于5‘-O-氨基甲酰腺苷(AMS)设计了一系列抑制非核糖体肽合成酶(NRPS)中腺化(A)结构域的紧密结合的抑制剂。我们最近开发了一种针对A结构域的亲和探针，可用于特异性分离和鉴定NRPS模块。我们的合成探针是l-Phe-AMS(L-Phe-AMS-Biotin)的生物素化变体，选择性地靶向NRPS模块中的A结构域，这些模块识别l-Phe并将其转化为整个蛋白质组中的氨基酰腺苷。在本章中，我们描述了l-Phe-AMS-Biotin的设计和合成，并总结了我们为使用该探针开发一系列特定浓缩NRPS模块的协议所做的工作。
A series of inhibitors have been designed based on 5′-O-sulfamoyl adenosine (AMS) that display tight binding characteristics towards the inhibition of adenylation (A) domains in nonribosomal peptide synthetases (NRPSs). We recently developed an affinity probe for A domains that could be used to facilitate the specific isolation and identification of NRPS modules. Our synthetic probe, which is a biotinylated variant ofl-Phe-AMS (l-Phe-AMS-biotin), selectively targets the A domains in NRPS modules that recognize and convertl-Phe to an aminoacyl adenylate in whole proteomes. In this chapter, we describe the design and synthesis ofl-Phe-AMS-biotin and provide a summary of our work towards the development of a series of protocols for the specific enrichment of NRPS modules using this probe.