Department of Biotechnology, Delft University of Technology, Delft, The Netherlands.
Winogradsky Institute of Microbiology, Research Centre of Biotechnology, Russian Academy of Sciences, Moscow, Russia.
ISME J. 2023 Oct;17(10):1639-1648. doi: 10.1038/s41396-023-01473-2. Epub 2023 Jul 13.
Dissimilatory nitrate reduction to ammonia (DNRA) is a common biochemical process in the nitrogen cycle in natural and man-made habitats, but its significance in wastewater treatment plants is not well understood. Several ammonifying Trichlorobacter strains (former Geobacter) were previously enriched from activated sludge in nitrate-limited chemostats with acetate as electron (e) donor, demonstrating their presence in these systems. Here, we isolated and characterized the new species Trichlorobacter ammonificans strain G1 using a combination of low redox potential and copper-depleted conditions. This allowed purification of this DNRA organism from competing denitrifiers. T. ammonificans is an extremely specialized ammonifier, actively growing only with acetate as e-donor and carbon source and nitrate as e-acceptor, but H can be used as an additional e-donor. The genome of G1 does not encode the classical ammonifying modules NrfAH/NrfABCD. Instead, we identified a locus encoding a periplasmic nitrate reductase immediately followed by an octaheme cytochrome c that is conserved in many Geobacteraceae species. We purified this octaheme cytochrome c protein (TaNiR), which is a highly active dissimilatory ammonifying nitrite reductase loosely associated with the cytoplasmic membrane. It presumably interacts with two ferredoxin subunits (NapGH) that donate electrons from the menaquinol pool to the periplasmic nitrate reductase (NapAB) and TaNiR. Thus, the Nap-TaNiR complex represents a novel type of highly functional DNRA module. Our results indicate that DNRA catalyzed by octaheme nitrite reductases is a metabolic feature of many Geobacteraceae, representing important community members in various anaerobic systems, such as rice paddy soil and wastewater treatment facilities.
异化硝酸盐还原为氨（DNRA）是自然和人工生境氮循环中的一种常见生化过程，但它在废水处理厂中的意义尚未得到很好的理解。先前已经从硝酸盐限制的恒化器中用乙酸作为电子（e）供体从活性污泥中富集了几种产氨三氯杆菌（前称为 Geobacter）菌株，证明了它们在这些系统中的存在。在这里，我们使用低氧化还原电位和缺铜条件的组合，从竞争的反硝化菌中分离和表征了新物种三氯杆菌属氨化菌 G1。这使得可以从竞争的反硝化菌中纯化这种 DNRA 生物。T. ammonificans 是一种极其特化的氨化菌，仅在乙酸作为电子供体和碳源以及硝酸盐作为电子受体的情况下才能积极生长，但 H 可以用作额外的电子供体。G1 的基因组不编码经典的氨化模块 NrfAH/NrfABCD。相反，我们鉴定了一个编码周质硝酸盐还原酶的基因座，该基因座紧随其后是一个八血红素细胞色素 c，该基因在许多 Geobacteraceae 物种中保守。我们纯化了这种八血红素细胞色素 c 蛋白（TaNiR），它是一种高度活跃的异化氨化亚硝酸盐还原酶，与细胞质膜松散结合。它可能与两个铁氧还蛋白亚基（NapGH）相互作用，从menaquinol 池向周质硝酸盐还原酶（NapAB）和 TaNiR 传递电子。因此，Nap-TaNiR 复合物代表了一种新型的高度功能化的 DNRA 模块。我们的结果表明，八血红素亚硝酸盐还原酶催化的 DNRA 是许多 Geobacteraceae 的代谢特征，代表了各种厌氧系统（如稻田土壤和废水处理设施）中的重要群落成员。
