Hydrolase CehA and Monooxygenase CfdC Are Responsible for Carbofuran Degradation in Sphingomonas sp Strain CDS-1
Hydrolase CehA and Monooxygenase CfdC Are Responsible for Carbofuran Degradation in Sphingomonas sp Strain CDS-1
复制标题
水解酶 CehA 和单加氧酶 CfdC 负责鞘氨醇单胞菌菌株 CDS-1 中克百威的降解
DOI：
10.1128/aem.00805-18
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发表时间：
2018
影响因子：
4.4
通讯作者：
Hong Q
中科院分区：
文献类型：
--
作者：
Yan Xin;Jin Wen;Wu Guang;Jiang Wankui;Yang Zhangong;Ji Junbin;Qiu Jiguo;He Jian;Jiang Ji;ong;Hong Qing;He Jian;Yan Xin;Hong Q
呋喃丹是一种广谱内吸性杀虫剂，已广泛使用约 50 年。多种克百威降解细菌已被描述，其中鞘氨醇单胞菌表现出非凡的分解代谢克百威的能力；其他细菌只能将呋喃丹转化为呋喃丹苯酚，而所有呋喃丹降解鞘氨醇单胞菌都可以降解呋喃丹和呋喃丹苯酚。然而，呋喃丹在鞘氨醇单胞菌中分解代谢的遗传基础尚未得到很好的阐明。在这项工作中，我们对鞘氨醇单胞菌的基因组草案进行了测序。菌株 CDS-1 可以转化呋喃丹和呋喃丹苯酚，但不能在它们上生长。基于克百威降解代谢相关基因在克百威降解鞘氨醇单胞菌中高度保守的假设，从菌株 CDS-1 和另一种鞘氨醇单胞菌 Novosphingobium sp 之间具有 ≥ 95% 核酸相似性的 84 个开放阅读框 (ORF) 中预测出两个这样的基因，cehACDS-1 和 cfdCCDS-1。菌株KN65.2能够矿化克百威的苯环。基因敲除、基因互补、异源表达和酶学实验结果表明，菌株CDS-1中cehACDS-1和cfdCCDS-1分别负责克百威和克百威苯酚的转化。 CehACDS-1 将克百威水解为克百威苯酚。 CfdCCDS-1 是一种还原型黄素单核苷酸 (FMNH2) 或还原型黄素腺嘌呤二核苷酸 (FADH2) 依赖性单加氧酶，在 NADH、FMN/FAD 和还原酶 CfdX 存在的情况下，可使苯环上的克百威苯酚羟基化。值得注意的是，我们发现先前被证明对克百威没有活性的西维因水解酶CehAAC100实际上可以将克百威转化为克百威苯酚，尽管活性非常低。 重要性由于克百威在过去50年的广泛使用，细菌已经进化出分解代谢途径来矿化这种杀虫剂，这在消除环境中克百威残留方面发挥着重要作用。这项研究揭示了鞘氨醇单胞菌中克百威降解的遗传决定因素。菌株CDS-1。我们推测，密切的同源物cehA和cfdC在其他克百威降解鞘氨醇单胞菌中高度保守，并且发挥与此处描述的相同的作用。这些研究结果加深了我们对呋喃丹微生物降解机制的认识，为更好地利用微生物修复呋喃丹污染奠定了基础。
Carbofuran, a broad-spectrum systemic insecticide, has been extensively used for approximately 50 years. Diverse carbofuran-degrading bacteria have been described, among which sphingomonads have exhibited an extraordinary ability to catabolize carbofuran; other bacteria can only convert carbofuran to carbofuran phenol, while all carbofuran-degrading sphingomonads can degrade both carbofuran and carbofuran phenol. However, the genetic basis of carbofuran catabolism in sphingomonads has not been well elucidated. In this work, we sequenced the draft genome of Sphingomonas sp. strain CDS-1 that can transform both carbofuran and carbofuran phenol but fails to grow on them. On the basis of the hypothesis that the genes involved in carbofuran catabolism are highly conserved among carbofuran-degrading sphingomonads, two such genes,cehACDS-1andcfdCCDS-1, were predicted from the 84 open reading frames (ORFs) that share ≥95% nucleic acid similarities between strain CDS-1 and another sphingomonad Novosphingobium sp. strain KN65.2 that is able to mineralize the benzene ring of carbofuran. The results of the gene knockout, genetic complementation, heterologous expression, and enzymatic experiments reveal thatcehACDS-1andcfdCCDS-1are responsible for the conversion of carbofuran and carbofuran phenol, respectively, in strain CDS-1. CehACDS-1hydrolyzes carbofuran to carbofuran phenol. CfdCCDS-1, a reduced flavin mononucleotide (FMNH2)- or reduced flavin adenine dinucleotide (FADH2)-dependent monooxygenase, hydroxylates carbofuran phenol at the benzene ring in the presence of NADH, FMN/FAD, and the reductase CfdX. It is worth noting that we found that carbaryl hydrolase CehAAC100, which was previously demonstrated to have no activity toward carbofuran, can actually convert carbofuran to carbofuran phenol, albeit with very low activity.IMPORTANCEDue to the extensive use of carbofuran over the past 50 years, bacteria have evolved catabolic pathways to mineralize this insecticide, which plays an important role in eliminating carbofuran residue in the environment. This study revealed the genetic determinants of carbofuran degradation in Sphingomonas sp. strain CDS-1. We speculate that the close homologuescehAandcfdCare highly conserved among other carbofuran-degrading sphingomonads and play the same roles as those described here. These findings deepen our understanding of the microbial degradation mechanism of carbofuran and lay a foundation for the better use of microbes to remediate carbofuran contamination.