Phytodetoxification of the explosive 2,4,6-trinitrotoluene

爆炸物 2,4,6-三硝基甲苯的植物解毒

• 批准号: BB/P005713/1
• 负责人: Neil Bruce
• 金额: $ 63.81万
• 依托单位: University of York
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FP005713%2F1
• 关键词: Phytodetoxification; explosive; trinitrotoluene

The explosive 2,4,6-trinitrotoluene (TNT) has become an extensive global pollutant over the last 100 years and there are mounting concerns over the toxicity of TNT to biological systems. During World War I and II the toxic effects of TNT were discovered during large scale production, with 475 fatalities and over 17,000 TNT poisoning cases reported at manufacturing facilities. TNT has been shown to severely impact the diversity of soil microbial communities and the establishment of vegetation. In the U.S. alone it is estimated that some 10 million hectares of military land is contaminated with munitions constituents. Unlike similar situations where the environment has become contaminated with toxic agrochemicals and their use subsequently banned, the huge demand for military explosives means that TNT will continue to be manufactured and used globally on a massive scale for the foreseeable future.Because of the scale of explosives pollution, particularly on military training ranges, there is considerable interest in developing plant based remediation strategies. Plants offer a low cost sustainable solution to containing and remediating explosives pollution. However, a fundamental understanding of the phytotoxicity of explosives, and the enzyme systems plants use to detoxify these compounds, and the rate-limiting steps, are required to enable the development of robust plant systems to contain and remediate explosives pollution effectively in situ. In plants, the majority of TNT remains in the roots, where it inhibits growth and development reducing whole plant biomass. We have recently discovered that the mitochondria- and plastid-targeted enzyme monodehydroascorbate reductase 6 (MDHAR6) reduces TNT by one electron, forming a nitro radical which reacts with atmospheric oxygen, generating highly reactive superoxide. This futile catalytic cycle only requires catalytic quantities of TNT to continuously generate damaging reactive oxygen species in the mitochondria. We have demonstrated that mutants in MDHAR6 have dramatically enhanced TNT tolerance, and we propose that this reaction accounts almost entirely for TNT toxicity in plants. The major goal of this research programme is to rigorously and quantitatively establish the fate and effects of TNT on plants. In order to achieve this objective we propose to study mechanisms of TNT toxicity and fully elucidate TNT induced detoxification pathways that include glucosylation, glutathionylation and oxidative activity by cytochromes P450. The fate of the TNT metabolites produced by these enzymes will then be established. We have previously demonstrated that the TNT active glutathione transferase GST-U25 results in the removal of a nitro group which could render the aromatic ring more amenable to biodegradation. We now have detailed structural information on GST-U25 that will allow us to engineer and improve the specificity and activity towards TNT. We hope to use the knowledge gained from this study to develop improved plant systems that will clean up polluted sites and prevent explosives pollution from contaminating water sources.

在过去的100年中，爆炸性的2,4,6-三硝基苯甲苯（TNT）已成为广泛的全球污染物，并且对TNT对生物系统的毒性存在持久的担忧。在第一次世界大战和II期间，在大规模生产中发现了TNT的毒性作用，在制造设施中报告了475例死亡和17,000多个TNT中毒病例。 TNT已被证明会严重影响土壤微生物群落的多样性和植被的建立。仅在美国，据估计，大约1000万公顷的军事土地被弹药选民污染。与类似的情况不同，环境已被有毒农化学污染及其随后被禁止的使用，对军事爆炸物的巨大需求意味着，TNT将继续在可预见的未来进行大规模制造和在全球范围内使用，因为爆炸物污染的规模是爆炸性培训范围，尤其是对工厂的兴趣，因此对工厂进行了基于工厂的兴趣。植物为包含和修复炸药污染提供了低成本的可持续解决方案。然而，需要对爆炸物的植物毒性的基本了解，并且需要使用酶系统来对这些化合物进行排毒，并且需要限制速率的步骤，以使能够开发强大的植物系统以遏制和修复爆炸物污染，从而有效地进行爆炸性污染。在植物中，大多数TNT仍然存在于根部，它抑制了生长和发育，从而减少了整个植物生物量。我们最近发现，线粒体和塑靶靶标的酶单羟苯甲酸盐还原酶6（MDHAR6）通过一个电子降低TNT，形成一种硝基自由基，形成了一种与大气中的氧反应，从而产生了高度反应的超级氧化物。这种徒劳的催化循环仅需要催化量的TNT即可在线粒体中连续产生破坏性的活性氧。我们已经证明，MDHAR6中的突变体具有显着增强的TNT耐受性，我们建议该反应几乎完全占植物中TNT的毒性。该研究计划的主要目标是严格，定量地确定TNT对植物的命运和影响。为了实现这一目标，我们建议研究TNT毒性的机制，并完全阐明TNT诱导的解毒途径，包括细胞色素P450在内的葡萄糖基化，谷胱甘肽化和氧化活性。然后将确定这些酶产生的TNT代谢产物的命运。我们先前已经证明，TNT活性谷胱甘肽转移酶GST-U25导致去除硝基基团，这可能使芳族环更适合生物降解。现在，我们在GST-U25上拥有详细的结构信息，这些信息将使我们能够设计并改善对TNT的特殊性和活动。我们希望利用这项研究所获得的知识来开发改进的植物系统，这些系统将清理污染的地点并防止爆炸物污染污染水源。

项目成果

A cofactor consumption screen identifies promising NfsB family nitroreductases for dinitrotoluene remediation.

辅因子消耗筛选确定了有前景的 NfsB 家族硝基还原酶用于二硝基甲苯修复。

• DOI: 10.1007/s10529-019-02716-z
• 发表时间: 2019
• 期刊: Biotechnology letters
• 影响因子: 2.7
• 作者: Williams EM