Identification and functional study of an iif2 gene cluster for indole degradation in Burkholderia sp. IDO3
Identification and functional study of an iif2 gene cluster for indole degradation in Burkholderia sp. IDO3
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伯克霍尔德氏菌 iif2 吲哚降解基因簇的鉴定和功能研究。
DOI：
10.1016/j.ibiod.2019.04.011
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发表时间：
2019-08
影响因子：
4.8
通讯作者：
Sun Yeqing
中科院分区：
文献类型：
--
作者：
Ma Qiao;Yang Bingyu;Qu Hui;Gao Zhen;Qu Yuanyuan;Sun Yeqing
伯克霍尔德里亚斯普。ID3是从活性污泥中分离到的一株吲哚降解菌。先前的基因组克隆文库分析确定了菌株ID03中吲哚代谢的aniif1基因簇。为了进一步探索吲哚降解的潜在机制，对菌株ID3的全基因组进行了测序(8,003,806bp)。该基因组包含3条环状染色体和1个质粒，预测了7550个基因。有趣的是，除了3号染色体上的iif1外，生物信息学分析还发现了1号染色体上的第二个吲哚加氧酶基因簇iif2。这两个基因簇都对吲哚有上调作用。在大肠杆菌BL21(DE3)中异源表达的ifC1D1和diifC2D2表明，这些基因能够将吲哚氧化成蓝色。基因敲除分析为C2是否在吲哚代谢中发挥关键作用提供了额外的证据。此外，我们在他们的iif2簇中发现了一个新的基因(IifF)。该基因被证明编码一种吲哚水解酶。表达IifF。Coli，获得了纯化的组氨酸标记酶制剂。IifF以Kmof4.4 ± 0.7 μM和Kcatof95.5 ± 4 S−1将吲哚转化为板蓝酸酯。这是首次研究表明吲哚可以被两个Iif基因簇降解，并且Isatin水解酶参与了吲哚代谢，加深了我们对吲哚代谢过程的理解。
Burkholderiasp. IDO3 is an indole-degrading bacterium isolated from activated sludge. A previous genomic clone library assay identified aniif1gene cluster for indole metabolism in strain IDO3. To further explore the underlying indole degradation mechanisms, the complete genome of strain IDO3 was sequenced (8,003,806 bp). The genome contained three circular chromosomes and one plasmid, and 7550 genes were predicted. Interestingly, in addition toiif1on chromosome 3, bioinformatic analyses identified a second indole oxygenase gene cluster,iif2, on chromosome 1. Bothiifclusters were up-regulated in response to indole. Heterologous expression ofiifC1D1andiifC2D2inEscherichia coliBL21(DE3) demonstrated that these genes were capable of oxidizing indole to indigo. Gene knockout assays provided additional evidence thatiifC2played crucial roles in indole metabolism. In addition, we identified a novel gene (iifF) in theiif2cluster. This gene was shown to encode an isatin hydrolase. IifF was expressed inE. coli, and a purified his-tagged enzyme preparation was obtained. IifF converted isatin to isatinate withKmof 4.4 ± 0.7 μM andkcatof 95.5 ± 4 s−1. This is the first study to show that indole can be degraded by twoiifgene clusters, and that isatin hydrolase is involved in indole metabolism, improving our understanding of indole metabolic processes.
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