Crystal structure of the nucleoid-associated protein Fis (PA4853) from Pseudomonas aeruginosa.
Crystal structure of the nucleoid-associated protein Fis (PA4853) from Pseudomonas aeruginosa.
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DOI：
10.1107/s2053230x20005427
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发表时间：
2020-04
期刊：
影响因子：
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通讯作者：
Juan Zhou;Zengqiang Gao;Heng Zhang;Yuhui Dong
中科院分区：
文献类型：
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作者：
Juan Zhou;Zengqiang Gao;Heng Zhang;Yuhui Dong
倒置刺激因子(FIS)是一种多功能的细菌类核相关蛋白，可以直接结合和弯曲DNA来影响DNA的拓扑结构。它还在调节细菌毒力因子和优化细菌对各种环境的适应方面发挥关键作用。最近发现，来自铜绿假单胞菌的FIS(PA4853，简称PaFis)是通过调节III型分泌系统(T3SS)基因的表达而产生毒力的。PaFis可以与exsA的启动子区域特异性结合，调控其表达，在exsB到exsA的转录延伸中起着至关重要的作用。本文报道了由四股螺旋束形成同源二聚体的PaFis的晶体结构。PaFis与研究较多的大肠杆菌Fis(EcFis)在结构上有显著的相似性，包括N端柔性环和C端螺旋-转角-螺旋(HTH)基序。然而，在ECFis的N-末端环中，Hin催化的DNA反转的关键残基在PaFis中并不保守，其确切作用还需要进一步的研究。凝胶电泳迁移率改变分析表明，PaFis能有效结合exsA启动子区域。基于结构的突变表明，HTH基序中的几个保守的碱性残基在DNA结合中起着至关重要的作用。这些结构和生化研究可能有助于理解PaFis在调节T3SS表达和毒力中的作用。
Factor for inversion stimulation (Fis) is a versatile bacterial nucleoid-associated protein that can directly bind and bend DNA to influence DNA topology. It also plays crucial roles in regulating bacterial virulence factors and in optimizing bacterial adaptation to various environments. Fis from Pseudomonas aeruginosa (PA4853, referred to as PaFis) has recently been found to be required for virulence by regulating the expression of type III secretion system (T3SS) genes. PaFis can specifically bind to the promoter region of exsA, which functions as a T3SS master regulator, to regulate its expression and plays an essential role in transcription elongation from exsB to exsA. Here, the crystal structure of PaFis, which is composed of a four-helix bundle and forms a homodimer, is reported. PaFis shows remarkable structural similarities to the well studied Escherichia coli Fis (EcFis), including an N-terminal flexible loop and a C-terminal helix-turn-helix (HTH) motif. However, the critical residues for Hin-catalyzed DNA inversion in the N-terminal loop of EcFis are not conserved in PaFis and further studies are required to investigate its exact role. A gel-electrophoresis mobility-shift assay showed that PaFis can efficiently bind to the promoter region of exsA. Structure-based mutagenesis revealed that several conserved basic residues in the HTH motif play essential roles in DNA binding. These structural and biochemical studies may help in understanding the role of PaFis in the regulation of T3SS expression and in virulence.