二价金属离子与YycFN相互作用的NMR研究

doi:10.11938/cjmr20160107

摘要/Abstract

摘要：

YycGF最早发现于枯草芽孢杆菌,是与细胞存活密切相关的双组分信号转导系统,存在于少量低鸟嘌呤和胞嘧啶(G+C)含量的革兰氏阳性菌中,包括金黄色葡萄球菌和肺炎链球菌等人类病原菌.在外界环境刺激下,胞膜上的组氨酸激酶YycG通过自身组氨酸磷酸化活化,将磷酸根转移至反应调节蛋白YycF的N端调节区(YycF_N)使之磷酸化,调控下游基因表达,实现特定细胞应答反应.二价金属离子在双组分信号转导系统反应调节蛋白的磷酸化过程中起着非常关键的作用,但它们与YycF_N相互作用的机制尚不清楚.该文利用液体核磁共振(NMR)方法研究了Ca²⁺、Mg²⁺两种离子与YycF_N的相互作用,对详细的相互作用界面进行了分析,并计算了Ca²⁺、Mg²⁺与YycF_N的解离常数(K_d).发现金属离子的关键作用位点是Asp9、Asp16和Asp53等残基,蛋白的整体构象也发生了一定变化,为阐明二价金属离子在反应调节蛋白信号转导过程中的作用机制提供了重要线索.

关键词: 核磁共振(NMR), 金属离子, 相互作用, YycF_N

Abstract:

YycGF, originally identified in Bacillus subtilis, is recognized as a crucial two-component signal transduction system closely associated with cell viability. It is highly conserved in low G+C Gram-positive bacteria, including Staphylococcus aureus, Streptococcus pneumoniae and other human pathogens. The histidine kinase (HK) YycG senses extracellular or intracellular signals and phosphorylates its cognate response regulator (RR) YycF, which in turn recognizes sequence specific regions on the bacterial chromosome and regulates the expression of certain genes. The presence of a divalent metal ion is essential for phosphoryl group transfer. Here, we presented a metal ion binding study of the response regulator YycF (YycF_N) from Bacillus subtilis using NMR spectroscopy. The metal ions Ca²⁺ and Mg²⁺ induced severe chemical shift changes in YycF backbone resonances, involving mainly the Asp9, Asp16 and Asp53 residues. Furthermore, the binding affinities of Ca²⁺ and Mg²⁺ with YycF_N were compared. The results provide important clues for understanding the conformational change of YycF_N upon metal ion binding before its phosphorylation.

Key words: NMR, metal ions, interaction, YycF_N

中图分类号:

O482.53

刘婷, 刘买利, 姜凌. 二价金属离子与YycF_N相互作用的NMR研究[J]. 波谱学杂志, 2016, 33(1): 77-88.

LIU Ting, LIU Mai-li, JIANG Ling. Divalent Metal Ion Binding to the Response Regulator YycF_N Studied by NMR Spectroscopy[J]. Chinese Journal of Magnetic Resonance, 2016, 33(1): 77-88.

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