All diazotrophic bacteria and archaea isolated so far utilise a nitrogenase enzyme-containing molybdenum in the active site co-factor to fix atmospheric dinitrogen to ammonia. However, in addition to the Mo-dependent nitrogenase, some nitrogen-fixing prokaryotes also express genetically distinct alternative nitrogenase isoenzymes, namely the V-dependent and Fe-only nitrogenases, respectively. Nitrogenase isoenzymes are expressed hierarchically according to metal availability and catalytic efficiency. In proteobacteria, this hierarchy is maintained via stringent transcriptional regulation of gene clusters by dedicated bacterial enhancer-binding proteins (bEBPs). The model diazotroph Azotobacter vinelandii contains two paralogs of the vanadium nitrogenase activator VnfA (henceforth, VnfA1), designated VnfA2 and VnfA3, with unknown functions. Here we demonstrate that the VnfA1 and VnfA3 bEBPs bind to the same target promoters in the Azotobacter vinelandii genome and co-activate a subset of genes in the absence of V, including the structural genes for the Fe-only nitrogenase. Co-activation is inhibited by the presence of V and is dependent on an accessory protein VnfZ that is co-expressed with VnfA3. Our studies uncover a plethora of interactions between bEBPs required for nitrogen fixation, revealing the unprecedented potential for fine-tuning the expression of alternative nitrogenases in response to metal availability.

中文翻译：

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旁系同源细菌增强子结合蛋白之间的相互作用使Azotobacter vinelandii中替代固氮酶的金属依赖性调节成为可能

迄今为止分离的所有固氮细菌和古细菌都利用活性位点辅因子中含固氮酶的钼将大气中的二氮固定为氨。然而，除了 Mo 依赖性固氮酶外，一些固氮原核生物还表达遗传上不同的替代固氮酶同工酶，即 V 依赖性固氮酶和仅 Fe 固氮酶。固氮酶同工酶根据金属可用性和催化效率分级表达。在变形菌中，这种层次结构是通过专门的细菌增强子结合蛋白 (bEBP) 对基因簇的严格转录调节来维持的。模型固氮菌Azotobacter vinelandii含有钒固氮酶激活剂 VnfA（以下简称 VnfA1）的两个旁系同源物，命名为 VnfA2 和 VnfA3，功能未知。在这里，我们证明 VnfA1 和 VnfA3 bEBP 与Azotobacter vinelandii基因组中的相同靶启动子结合，并在没有 V 的情况下共同激活一部分基因，包括仅 Fe 固氮酶的结构基因。共激活被 V 的存在抑制并且依赖于与 VnfA3 共表达的辅助蛋白 VnfZ。我们的研究揭示了固氮所需的 bEBP 之间的大量相互作用，揭示了根据金属可用性微调替代固氮酶表达的前所未有的潜力。