Interfaculty Institute of Microbiology and Infection Medicine, Department of Organismic Interactions, Eberhard Karls University of Tübingen, Auf der Morgenstelle 28, 72076, Tübingen, Germany.
Sci Rep. 2019 Dec 12;9(1):18985. doi: 10.1038/s41598-019-55495-y.
PII proteins comprise an ancient superfamily of signal transduction proteins, widely distributed among all domains of life. In general, PII proteins measure and integrate the current carbon/nitrogen/energy status of the cell through interdependent binding of ATP, ADP and 2-oxogluterate. In response to effector molecule binding, PII proteins interact with various PII-receptors to tune central carbon- and nitrogen metabolism. In cyanobacteria, PII regulates, among others, the key enzyme for nitrogen-storage, N-acetyl-glutamate kinase (NAGK), and the co-activator of the global nitrogen-trascription factor NtcA, the PII-interacting protein-X (PipX). One of the remarkable PII variants from Synechococcus elongatus PCC 7942 that yielded mechanistic insights in PII-NAGK interaction, is the NAGK-superactivating variant I86N. Here we studied its interaction with PipX. Another critical residue is Lys58, forming a salt-bridge with 2-oxoglutarate in a PII-ATP-2-oxoglutarate complex. Here, we show that Lys58 of PII protein is a key residue for mediating PII interactions. The K58N mutation not only causes the loss of 2-oxogluterate binding but also strongly impairs binding of ADP, NAGK and PipX. Remarkably, the exchange of the nearby Leu56 to Lys in the K58N variant partially compensates for the loss of K58. This study demonstrates the potential of creating custom tailored PII variants to modulate metabolism.
PII 蛋白构成了信号转导蛋白的古老超家族，广泛分布于所有生命领域。一般来说，PII 蛋白通过与 ATP、ADP 和 2-氧代戊二酸的相互依赖结合来测量和整合细胞的当前碳/氮/能量状态。响应效应分子结合，PII 蛋白与各种 PII 受体相互作用，调节中心碳和氮代谢。在蓝藻中，PII 调节氮储存的关键酶，N-乙酰-谷氨酸激酶（NAGK），以及全局氮转录因子 NtcA 的共激活子，PII 相互作用蛋白-X（PipX）。来自 Synechococcus elongatus PCC 7942 的一种显著的 PII 变体，它提供了关于 PII-NAGK 相互作用的机制见解，是 NAGK 超激活变体 I86N。在这里，我们研究了它与 PipX 的相互作用。另一个关键残基是 Lys58，它在 PII-ATP-2-氧代戊二酸复合物中与 2-氧代戊二酸形成盐桥。在这里，我们表明 PII 蛋白的 Lys58 是介导 PII 相互作用的关键残基。K58N 突变不仅导致 2-氧代戊二酸结合的丧失，而且强烈损害 ADP、NAGK 和 PipX 的结合。值得注意的是，在 K58N 变体中附近的 Leu56 交换为 Lys 部分补偿了 K58 的丧失。这项研究表明了创建定制 PII 变体来调节代谢的潜力。
