Activation and characterization of a cryptic gene cluster reveals a cyclization cascade for polycyclic tetramate macrolactams.
Activation and characterization of a cryptic gene cluster reveals a cyclization cascade for polycyclic tetramate macrolactams.
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DOI：
10.1039/c6sc03875a
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发表时间：
2017-02-01
期刊：
影响因子：
8.4
通讯作者：
Zhang C
中科院分区：
文献类型：
--
作者：
Saha S;Zhang W;Zhang G;Zhu Y;Chen Y;Liu W;Yuan C;Zhang Q;Zhang H;Zhang L;Zhang W;Zhang C
我们报道了在海洋来源的平坦链霉菌中PtM基因簇的激活，导致发现了六个新的PtM，即PATAMIES A-F。多环四酸大环内酰胺类(PTM)是一类由聚酮合成酶(PKS)/非核糖体肽合成酶(NRPS)混合途径合成的天然产物。PTM生物合成基因簇在细菌中是保守的，分布广泛，但大多数都是沉默的。在这篇文章中，我们报道了通过启动子工程和异源表达，激活了海洋来源的松香链霉菌SCSIO 02999中的Ptm基因簇，导致发现了6个新的Ptm，Pacamide A-F(11-16)，对几种人类癌细胞具有很强的细胞毒活性。体内基因阻断实验和体外生化分析表明，多环的形成是一个还原环化级联反应，反应依次产生5，5/5和5/5/6碳环系统，分别由植烯脱氢酶PtmB2，氧化还原酶PtmB1和乙醇脱氢酶PtmC催化。此外，PtmC还被证明是一种双功能环化酶，可以催化伊卡鲁霉素内五元环的形成。本研究提出了通过基因组挖掘发现更具生物活性的PTMS的可能性，并揭示了PTMS中5/5/6型碳环形成的一般机制。
We report the activation of a PTM gene cluster in marine-derived Streptomyces pactum, leading to the discovery of six new PTMs, the pactamides A-F. Polycyclic tetramate macrolactams (PTMs) are a growing class of natural products and are derived from a hybrid polyketide synthase (PKS)/non-ribosomal peptide synthetase (NRPS) pathway. PTM biosynthetic gene clusters are conserved and widely distributed in bacteria, however, most of them remain silent. Herein we report the activation of a PTM gene cluster in marine-derived Streptomyces pactum SCSIO 02999 by promoter engineering and heterologous expression, leading to the discovery of six new PTMs, pactamides A–F (11–16), with potent cytotoxic activity upon several human cancer cell lines. In vivo gene disruption experiments and in vitro biochemical assays reveal a reductive cyclization cascade for polycycle formation, with reactions sequentially generating the 5, 5/5 and 5/5/6 carbocyclic ring systems, catalysed by the phytoene dehydrogenase PtmB2, the oxidoreductase PtmB1, and the alcohol dehydrogenase PtmC, respectively. Furthermore, PtmC was demonstrated as a bifunctional cyclase for catalyzing the formation of the inner five-membered ring in ikarugamycin. This study suggests the possibility of finding more bioactive PTMs by genome mining and discloses a general mechanism for the formation of 5/5/6-type carbocyclic rings in PTMs.