GlnK Facilitates the Dynamic Regulation of Bacterial Nitrogen Assimilation.
GlnK Facilitates the Dynamic Regulation of Bacterial Nitrogen Assimilation.
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DOI：
10.1016/j.bpj.2017.04.012
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发表时间：
2017-05-23
影响因子：
3.4
通讯作者：
Barahona M
中科院分区：
文献类型：
--
作者：
Gosztolai A;Schumacher J;Behrends V;Bundy JG;Heydenreich F;Bennett MH;Buck M;Barahona M
大肠杆菌中的氨同化受两种类似蛋白(GlnB和GlnK)的调控，这两种蛋白协调与基因表达、运输蛋白和代谢途径的调节。然而，它们是如何联合调节谷氨酰胺合成酶的活性的，人们对此知之甚少。谷氨酰胺合成酶是氮吸收的关键酶。我们采用实验和理论相结合的方法，研究了GlnB和GlnK在氮素饥饿和上移过程中的动态作用。我们测量了体内代谢物、总蛋白和翻译后修饰蛋白的时间分辨浓度，并开发了一个简明的GlnB和GlnK生化模型，该模型包含了对活性和变构位点的竞争以及GlnK的功能隔离。该模型预测了谷氨酰胺合成酶、GlnB和GlnK在野生型和两种基因敲除中随时间变化的外源铵水平的反应。我们的结果表明，GlnK在富氮条件下受到严格调控，但在氨耗尽和饥饿时表达。这表明GlnK在饥饿后作为氮休克的缓冲作用，并在氮和碳代谢之间提供了进一步的功能联系。
Ammonium assimilation in Escherichia coli is regulated by two paralogous proteins (GlnB and GlnK), which orchestrate interactions with regulators of gene expression, transport proteins, and metabolic pathways. Yet how they conjointly modulate the activity of glutamine synthetase, the key enzyme for nitrogen assimilation, is poorly understood. We combine experiments and theory to study the dynamic roles of GlnB and GlnK during nitrogen starvation and upshift. We measure time-resolved in vivo concentrations of metabolites, total and posttranslationally modified proteins, and develop a concise biochemical model of GlnB and GlnK that incorporates competition for active and allosteric sites, as well as functional sequestration of GlnK. The model predicts the responses of glutamine synthetase, GlnB, and GlnK under time-varying external ammonium level in the wild-type and two genetic knock-outs. Our results show that GlnK is tightly regulated under nitrogen-rich conditions, yet it is expressed during ammonium run-out and starvation. This suggests a role for GlnK as a buffer of nitrogen shock after starvation, and provides a further functional link between nitrogen and carbon metabolisms.