Metabolic diversity in myxobacteria:: identification of the myxalamid and the stigmatellin biosynthetic gene cluster of Stigmatella aurantiaca Sg a15 and a combined polyketide-(poly)peptide gene cluster from the epothilone producing strain Sorangium cellulosum So ce90
Metabolic diversity in myxobacteria:: identification of the myxalamid and the stigmatellin biosynthetic gene cluster of Stigmatella aurantiaca Sg a15 and a combined polyketide-(poly)peptide gene cluster from the epothilone producing strain Sorangium cellulosum So ce90
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DOI：
10.1016/s0167-4781(99)00041-x
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发表时间：
1999-05-14
期刊：
影响因子：
--
通讯作者：
Müller, R
中科院分区：
文献类型：
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作者：
Beyer, S;Kunze, B;Müller, R
粘细菌菌株生产的聚酮（PKs）假定由I型聚酮合酶（PKS）的生物合成进行了分析。粘细菌还产生多种多肽（PP）和具有掺入的氨基酸的PK（“混合PK-PP”）。为了能够鉴定这些代谢物的生物合成基因簇，已经开发了基于PCR的方法来克隆来自这些生物体的PKS基因的酮合酶（KS）结构域。非核糖体型肽合成酶（NRPS）的保守区域也通过PCR扩增。将来自橙色柱头菌Sg a15的KS片段用于染色体基因失活实验，产生一系列突变体，包括不能产生柱头菌素和粘素酰胺的突变体。用纤维堆囊菌（SorangiumcellulosumSo ce 90）的NRPS片段和PKS片段鉴定与来自基因组文库的两种探针杂交的互补序列。NRPS和PKS片段都是从这些互补序列之一的相对较短的限制性片段克隆和测序的。这里描述的方法应该是非常有用的克隆和鉴定PKS，NRPS和混合PKS-NRPS从粘细菌一般，从而打开机会，利用这些细菌的生物化学多样性的基因工程和组合生物合成。(C)1999 Elsevier Science B. V.保留所有权利。
Myxobacterial strains producing polyketides (PKs) assumed to be biosynthesized by a type I polyketide synthase (PKS) were analysed. Myxobacteria also produce a variety of polypeptides (PP) and PKs with incorporated amino acids ('mixed PK-PP'). In order to be able to identify the biosynthetic gene clusters for these metabolites a PCR based approach has been developed to clone ketosynthase (KS) domains of PKS genes from these organisms. Conserved regions of peptide synthetases of the non-ribosomal type (NRPS) were also amplified via PCR. KS fragments from Stigmatella aurantiaca Sg a15 were used for chromosomal gene inactivation experiments resulting in a series of mutants including such that were unable to produce stigmatellins and myxalamids. A NRPS fragment and PKS fragments from Sorangium cellulosum So ce90 were used to identify cosmids hybridizing with both types of probes from a genomic library. Both a NRPS and a PKS fragment were cloned and sequenced from a relatively short restriction fragment of one of these cosmids. The method described here should be very useful to clone and identify PKS, NRPS and mixed PKS-NRPS from myxobacteria in general and thereby open opportunities to use the biochemical diversity of these bacteria for genetic engineering and combinatorial biosynthesis. (C) 1999 Elsevier Science B.V. All rights reserved.