Combinatorial polyketide biosynthesis by de novo design and rearrangement of modular polyketide synthase genes
Combinatorial polyketide biosynthesis by de novo design and rearrangement of modular polyketide synthase genes
复制标题
DOI：
10.1038/nbt1128
复制
发表时间：
2005-09-01
影响因子：
46.9
通讯作者：
Santi, DV
中科院分区：
文献类型：
--
作者：
Menzella, HG;Reid, R;Santi, DV
I型聚酮合成酶(PKS)基因由3-6kb长的模块组成，编码聚酮产物中2-碳单元的结构。个别PKS模块的改变或更换可能导致“非天然”天然产物的生物合成，但现有的技术是耗时的。在这里，我们描述了一种设计合成PKS基因的通用方法，其中通过一组重复的侧翼限制位点来促进模块和结构域的简易盒组装和互换。为了验证这种方法的可行性，我们从8个PKS簇中合成了14个模块，并将它们关联在154个双模组合中，这些组合跨越了超过150万个核苷酸的新的PKS基因序列。近一半的组合在大肠杆菌中成功地介导了聚酮的生物合成，所有单独的模块都参与了生产性的双峰组合。这项工作为聚酮的生产提供了一种真正的组合方法。
Type I polyketide synthase (PKS) genes consist of modules similar to 3-6 kb long, which encode the structures of 2-carbon units in polyketide products. Alteration or replacement of individual PKS modules can lead to the biosynthesis of 'unnatural' natural products but existing techniques for this are time consuming. Here we describe a generic approach to the design of synthetic PKS genes where facile cassette assembly and interchange of modules and domains are facilitated by a repeated set of flanking restriction sites. To test the feasibility of this approach, we synthesized 14 modules from eight PKS clusters and associated them in 154 bimodular combinations spanning over 1.5-million bp of novel PKS gene sequences. Nearly half the combinations successfully mediated the biosynthesis of a polyketide in Escherichia coli, and all individual modules participated in productive bimodular combinations. This work provides a truly combinatorial approach for the production of polyketides.