Expanding Small-Molecule Functional Metagenomics through Parallel Screening of Broad-Host-Range Cosmid Environmental DNA Libraries in Diverse Proteobacteria
Expanding Small-Molecule Functional Metagenomics through Parallel Screening of Broad-Host-Range Cosmid Environmental DNA Libraries in Diverse Proteobacteria
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DOI：
10.1128/aem.02169-09
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发表时间：
2010-03-01
影响因子：
4.4
通讯作者：
Brady, Sean F.
中科院分区：
文献类型：
--
作者：
Craig, Jeffrey W.;Chang, Fang-Yuan;Brady, Sean F.
未培养细菌基因组中编码的小分子生物合成多样性是利用功能元基因组学发现天然产物的一个有吸引力的目标。通常与小分子产生相关的表型，如抗生素、色素改变或菌落形态改变，很容易从排列的元基因组文库克隆的筛选中识别出来。然而，功能性元基因组筛选方法因其对异源表达宿主的内在依赖性而受到限制。为了增加在功能元基因组研究中发现的小分子生物合成多样性，我们报道了在六种不同的蛋白细菌中对广泛宿主范围的环境DNA文库的表型筛选：根癌农杆菌、伯克霍尔德氏菌、弧形杆菌、大肠杆菌、恶臭假单胞菌和变态罗氏杆菌。描述了在发色和抗菌活性克隆的发酵液提取物中发现的克隆特异性小分子，以及负责这些代谢物生物合成的遗传元件。这项研究中使用的宿主菌株提供了获得重叠最少的独特克隆集的途径，从而展示了通过使用多种不同的宿主物种赋予功能元基因组学的潜在优势。
The small-molecule biosynthetic diversity encoded within the genomes of uncultured bacteria is an attractive target for the discovery of natural products using functional metagenomics. Phenotypes commonly associated with the production of small molecules, such as antibiosis, altered pigmentation, or altered colony morphology, are easily identified from screens of arrayed metagenomic library clones. However, functional metagenomic screening methods are limited by their intrinsic dependence on a heterologous expression host. Toward the goal of increasing the small-molecule biosynthetic diversity found in functional metagenomic studies, we report the phenotypic screening of broad-host-range environmental DNA libraries in six different proteobacteria: Agrobacterium tumefaciens, Burkholderia graminis, Caulobacter vibrioides, Escherichia coli, Pseudomonas putida, and Ralstonia metallidurans. Clone-specific small molecules found in culture broth extracts from pigmented and antibacterially active clones, as well as the genetic elements responsible for the biosynthesis of these metabolites, are described. The host strains used in this investigation provided access to unique sets of clones showing minimal overlap, thus demonstrating the potential advantage conferred on functional metagenomics through the use of multiple diverse host species.