División de Bioquímica Y Biología Molecular, Departamento de Agroquímica Y Bioquímica, Universidad de Alicante, Carretera de San Vicente del Raspeig s/n, San Vicente del Raspeig, 03690, Alicante, Spain.
Departamento de Matemática Aplicada, Universidad de Alicante, Carretera de San Vicente del Raspeig s/n, San Vicente del Raspeig, 03690, Alicante, Spain.
Extremophiles. 2020 May;24(3):433-446. doi: 10.1007/s00792-020-01169-x. Epub 2020 Apr 16.
Glutamine synthetase is an essential enzyme in ammonium assimilation and glutamine biosynthesis. The Haloferax mediterranei genome has two other glnA-type genes (glnA2 and glnA3) in addition to the glutamine synthetase gene glnA. To determine whether the glnA2 and glnA3 genes can replace glnA in nitrogen metabolism, we generated deletion mutants of glnA. The glnA deletion mutants could not be generated in a medium without glutamine, and thus, glnA is an essential gene in H. mediterranei. The glnA deletion mutant was achieved by adding 40 mM glutamine to the selective medium. This conditional HM26-ΔglnA mutant was characterised with different approaches in the presence of distinct nitrogen sources and nitrogen starvation. Transcriptomic analysis was performed to compare the expression profiles of the strains HM26-ΔglnA and HM26 under different growth conditions. The glnA deletion did not affect the expression of glnA2, glnA3 and nitrogen assimilation genes under nitrogen starvation. Moreover, the results showed that glnA, glnA2 and glnA3 were not expressed under the same conditions. These results indicated that glnA is an essential gene for H. mediterranei and, therefore, glnA2 and glnA3 cannot replace glnA in the conditions analysed.
谷氨酰胺合成酶是铵同化和谷氨酰胺生物合成的必需酶。除了谷氨酰胺合成酶基因 glnA 外，极端嗜盐古菌地中海盐菌基因组中还有另外两个 glnA 型基因（glnA2 和 glnA3）。为了确定 glnA2 和 glnA3 基因是否可以替代氮代谢中的 glnA，我们生成了 glnA 的缺失突变体。在没有谷氨酰胺的培养基中无法生成 glnA 缺失突变体，因此，glnA 是地中海盐菌中的必需基因。通过在选择培养基中添加 40 mM 谷氨酰胺来实现 glnA 缺失突变体。利用不同的氮源和氮饥饿条件，通过不同的方法对具有条件性 HM26-ΔglnA 突变体的特性进行了研究。通过比较不同生长条件下菌株 HM26-ΔglnA 和 HM26 的转录组分析，来比较它们的表达谱。在氮饥饿条件下，glnA 缺失不影响 glnA2、glnA3 和氮同化基因的表达。此外，结果表明 glnA、glnA2 和 glnA3 在相同条件下不表达。这些结果表明，glnA 是地中海盐菌的必需基因，因此，在分析条件下，glnA2 和 glnA3 不能替代 glnA。
