Cloning and heterologous expression of the macrotetrolide biosynthetic gene cluster revealed a novel polyketide synthase that lacks an acyl carrier protein
Cloning and heterologous expression of the macrotetrolide biosynthetic gene cluster revealed a novel polyketide synthase that lacks an acyl carrier protein
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DOI：
10.1021/ja0100827
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发表时间：
2001-04-11
影响因子：
15
通讯作者：
Shen, B
中科院分区：
文献类型：
--
作者：
Kwon, HJ;Smith, WC;Shen, B
大环四内酯是一类环状聚醚，由四个对映异构体九乳酸（1）或其同系物（+）（-）（+）（-）-酯键1组成，具有广谱生物活性，2包括抗菌、抗真菌、抗肿瘤和免疫抑制活性。对灰色链霉菌中大环内酯生物合成的研究已经明确确定它们是聚酮化合物来源。3，4特别感兴趣的是大环内酯生物合成涉及一对对映体特异性聚酮途径4c的提议（图1）。（a）（6 R，8 R）-或（6S，8 S）-2-甲基-6，8-二羟基壬-2E-烯酸（2）3d和（（）-13 a有效地掺入到nonactin（3）中，（B）从S中分离出（+）-和（-）-1及其二聚体3g。（c）催化从（-）-2及其同系物形成（-）-1及其同系物的NonS的遗传和生物化学表征。4 b，c这一提议的一个有趣的推论是合成一组对映体聚酮化合物，如2，推测是由一对聚酮化合物酶（PKS）。4c为了从羧酸前体制备2，后面的PKS必须调用（a）很少使用琥珀酸作为完整的四碳片段（C3-C6）和（B）衍生出三碳单元（C7-C9）从两个醋酸分子。我们克隆了大环内酯生物合成基因簇，并进行了异源表达，揭示了一种新的聚酮生物合成PKS，它由II型酮脂酰脱氢酶（KS）和酮还原酶（KRs）组成，但缺少酰基载体蛋白（ACP）。利用先前表征的nonR抗性基因作为探针，4a，c nonS（与nonR相邻的基因）的失活完全消除了大环内酯的产生，证实了克隆的DNA编码大环内酯的生物合成。4c核苷酸
The macrotetrolides are a family of cyclic polyethers composed of four molecules of enantiomeric nonactic acid (1) or its homologues in a (+)(-)(+)(-)-ester linkage1 and exhibit a broad spectrum of biological activities, 2 ranging from antibacterial, antifungal, antitumor, to immunosuppressive activity. Studies of macrotetrolide biosynthesis in Streptomyces griseus have unambiguously established that they are of polyketide origin. 3, 4 Of special interest is the proposal that macrotetrolide biosynthesis involves a pair of enantiospecific polyketide pathways4c (Figure 1). This idea is supported by (a) the efficient incorporation of (6R, 8R)-or (6S, 8S)-2-methyl-6, 8-dihydroxynon-2E-enoic acid (2) 3d and (()-13a into nonactin (3),(b) the isolation of both (+)-and (-)-1 and their dimers3g from S. griseus fermentation, and (c) the genetic and biochemical characterization of NonS that catalyzes formation of (-)-1 and its homologues from (-)-2 and its homologues. 4b, c An intriguing corollary of this proposal is the synthesis of a set of enantiomeric polyketides, such as 2, presumably by a pair of polyketide synthases (PKSs). 4c To make 2 from the carboxylic acid precursors, the latter PKSs must have invoked (a) the rare use of succinate as an intact four-carbon fragment (C3-C6) and (b) the derivation of a three-carbon unit (C7-C9) from two molecules of acetate. We now report the cloning and heterologous expression of the macrotetrolide biosynthetic gene cluster, revealing a novel type of PKS for polyketide biosynthesis that consists of type II ketoacyl synthases (KSs) and ketoreductases (KRs) but lacks acyl carrier protein (ACP).A 55-kb contiguous DNA region was cloned from S. griseus DSM40695 using the previously characterized nonR resistance gene as a probe. 4a, c Inactivation of nonS, a gene adjacent to nonR, completely abolished macrotetrolide production, confirming that the cloned DNA encodes macrotetrolide biosynthesis. 4c Nucleotide