Heterologous expression of nitrogenase has been actively pursued because of the far-reaching impact of this enzyme on agriculture, energy and the environment. However, isolation of an active two-component, metallocentre-containing nitrogenase from a non-diazotrophic host has yet to be accomplished. Here we report the heterologous synthesis of an active molybdenum-nitrogenase by combining genes from Azotobacter vinelandii and Methanosarcina acetivorans in Escherichia coli. Metal, activity and electron paramagnetic resonance analyses demonstrate the integrity of the metallocentres in the purified nitrogenase enzyme; whereas growth, nanoscale secondary ion mass spectrometry and nuclear magnetic resonance experiments illustrate diazotrophic growth and ¹⁵N enrichment by the E. coli expression strain, and accumulation of extracellular ammonia upon deletion of the ammonia transporter that permits incorporation of thus-generated nitrogen into the cellular mass of a non-diazotrophic E. coli strain. As such, this study provides a crucial prototype system that could be optimized/modified to enable future transgenic expression and biotechnological adaptations of nitrogenase.

由于固氮酶对农业、能源和环境的深远影响，固氮酶的异源表达一直受到人们的积极关注。然而，从非固氮宿主中分离活性双组分、含金属中心的固氮酶尚未完成。在这里，我们报告了通过结合大肠杆菌中的固氮菌和乙酸甲烷八叠球菌的基因来异源合成活性钼固氮酶。金属、活性和电子顺磁共振分析证明了纯化固氮酶中金属中心的完整性；而生长、纳米级二次离子质谱和核磁共振实验说明了大肠杆菌表达菌株的固氮生长和¹⁵ N 富集，以及在氨转运蛋白缺失后细胞外氨的积累，从而允许将由此产生的氮掺入细胞中非固氮大肠杆菌菌株的质量。因此，这项研究提供了一个重要的原型系统，可以对其进行优化/修改，以实现未来的固氮酶转基因表达和生物技术适应。