Ishikawa Fumihiro, Kakeya Hideaki
Department of System Chemotherapy and Molecular Sciences, Division of Bioinformatics and Chemical Genomics, Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan.
Bioorg Med Chem Lett. 2014 Feb 1;24(3):865-9. doi: 10.1016/j.bmcl.2013.12.082. Epub 2013 Dec 25.
We targeted the development of an affinity probe for adenylation (A) domains that can facilitate enrichment, identification, and quantification of A domain-containing modules in nonribosomal peptide synthetase (NRPS)-polyketide synthase (PKS) hybrids and NRPSs. A 5'-O-sulfamoyladenosine (AMS) non-hydrolyzable analogue of adenosine monophosphate (AMP) has been reported as a scaffold for the design of inhibitors exhibiting tight binding of adenylation enzymes. Here we describe the application of an affinity probe for A domains. Our synthetic probe, a biotinylated L-Phe-AMS (L-Phe-AMS-biotin) specifically targets the A domains in NRPS modules that activates L-Phe to an aminoacyladenylate intermediate in both recombinant NRPS enzyme systems and whole proteomes.
我们致力于开发一种用于腺苷化（A）结构域的亲和探针，该探针能够促进非核糖体肽合成酶（NRPS）-聚酮合酶（PKS）杂合体和NRPS中含A结构域模块的富集、鉴定和定量。5'-O-氨磺酰腺苷（AMS），一种单磷酸腺苷（AMP）的不可水解类似物，已被报道作为设计与腺苷化酶紧密结合的抑制剂的支架。在此，我们描述了一种用于A结构域的亲和探针的应用。我们的合成探针，生物素化的L-苯丙氨酸-AMS（L-Phe-AMS-生物素），在重组NRPS酶系统和完整蛋白质组中，特异性靶向NRPS模块中的A结构域，该结构域将L-苯丙氨酸激活为氨酰腺苷酸中间体。
