Proteomics and Genetics for Identification of a Bacterial Antimonite Oxidase in Agrobacterium tumefaciens
Proteomics and Genetics for Identification of a Bacterial Antimonite Oxidase in Agrobacterium tumefaciens
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用于鉴定根癌农杆菌中细菌锑氧化酶的蛋白质组学和遗传学。
DOI：
10.1021/es506318b
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发表时间：
2015-05-19
影响因子：
11.4
通讯作者：
Wang, Gejiao
中科院分区：
文献类型：
--
作者：
Li, Jingxin;Wang, Qian;Wang, Gejiao
锑（Sb）及其化合物被美国环境保护局（USEPA, 1979）和欧盟（CEC, 1976）列为优先污染物。微生物氧化还原转化被认为是自然界中锑循环的重要组成部分；然而，这些过程的调控和所涉及的酶学尚不清楚。本研究通过对Sb（III）氧化菌农杆菌GW4的比较蛋白质组学和逆转录- pcr分析发现，氧化还原酶（anoA）广泛分布于微生物中，包括至少一些文献记载的能够氧化Sb（III）的微生物。anoA基因的缺失降低了Sb（III）抗性，并使Sb（III）氧化降低了约27%，而anoA补充菌株与野生型GW4相似，而GW4过表达anoA的菌株使Sb（III）氧化增加了约34%。添加Sb（III）上调anoA的表达，并将anoA克隆到大肠杆菌中，证明了这种活性的直接可转移性。经His-tag纯化的AnoA需要NADP（+）作为辅助因子，对Sb（III）的K(m)为64±10 μM， V（max）为150±7 nmol min(-1) mg（-1）。这项研究为了解微生物-锑相互作用的机制迈出了重要的第一步，并增强了我们对微生物如何参与自然界中锑的生物地球化学循环的理解。
Antimony (Sb) and its compounds are listed by the United States Environmental Protection Agency (USEPA, 1979) and the European Union (CEC, 1976) as a priority pollutant. Microbial redox transformations are presumed to be an important part of antimony cycling in nature; however, regulation of these processes and the enzymology involved are unknown. In this study, comparative proteomics and reverse transcriptase-PCR analysis of Sb(III)-oxidizing bacterium Agrobacterium tumefaciens GW4 revealed an oxidoreductase (anoA) is widely distributed in microorganisms, including at least some documented to be able to oxidize Sb(III). Deletion of the anoA gene reduced Sb(III) resistance and decreased Sb(III) oxidation by ∼27%, whereas the anoA complemented strain was similar to the wild type GW4 and a GW4 anoA overexpressing strain increased Sb(III) oxidation by ∼34%. Addition of Sb(III) up-regulated anoA expression and cloning anoA to Escherichia coli demonstrated direct transferability of this activity. A His-tag purified AnoA was found to require NADP(+) as cofactor, and exhibited a K(m) for Sb(III) of 64 ± 10 μM and a V(max) of 150 ± 7 nmol min(-1) mg(-1). This study contributes important initial steps toward a mechanistic understanding of microbe-antimony interactions and enhances our understanding of how microorganisms participate in antimony biogeochemical cycling in nature.