A mutagenic screen reveals NspS residues important for regulation of Vibrio cholerae biofilm formation.
A mutagenic screen reveals NspS residues important for regulation of Vibrio cholerae biofilm formation.
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诱变筛选揭示了 NspS 残基对于调节霍乱弧菌生物膜形成很重要。
DOI：
10.1099/mic.0.001023
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Kuhn,MistyL
中科院分区：
文献类型：
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作者：
Young,ErinC;Baumgartner,JacksonT;Karatan,Ece;Kuhn,MistyL
去亚精胺、亚精胺和精胺对人类肠道病原菌霍乱弧菌生物膜形成的部分调控作用。霍乱通过由质周蛋白、NspS和整体膜c-二gmp磷酸二酯酶MbaA组成的信号通路感知这些多胺。NspS和MbaA属于一类新的信号系统，由质周配体结合蛋白和含有GGDEF和EAL结构域的膜结合c-di-GMP磷酸二酯酶组成。在这一信号转导途径中，NspS被假设与外周质中的MbaA相互作用，调节其磷酸二酯酶活性。多胺与NspS结合可能改变这种相互作用，导致生物膜形成的激活或抑制取决于多胺。本研究的目的是确定对NspS功能重要的氨基酸。我们对该基因进行了随机诱变，鉴定了缺乏生物膜形成的突变克隆，确定了它们对去亚精胺的反应性，并将这些残基的位置映射到NspS同源模型上。单突变体聚集在NspS模型的两个裂片上，但大多数突变体出现在一个裂片上，该裂片在去亚精胺结合后似乎更具移动性。我们还在假设的配体结合位点上发现了残基，这些残基可能对去亚精胺与MbaA的结合和相互作用很重要。最终，我们的研究结果为霍乱这种新的信号通路提供了新的见解，并强调了参与转运与信号转导的质周结合蛋白之间的差异。
Biofilm formation in the human intestinal pathogenVibrio choleraeis in part regulated by norspermidine, spermidine and spermine.V. choleraesenses these polyamines through a signalling pathway consisting of the periplasmic protein, NspS, and the integral membrane c-di-GMP phosphodiesterase MbaA. NspS and MbaA belong to a proposed class of novel signalling systems composed of periplasmic ligand-binding proteins and membrane-bound c-di-GMP phosphodiesterases containing both GGDEF and EAL domains. In this signal transduction pathway, NspS is hypothesized to interact with MbaA in the periplasm to regulate its phosphodiesterase activity. Polyamine binding to NspS likely alters this interaction, leading to the activation or inhibition of biofilm formation depending on the polyamine. The purpose of this study was to determine the amino acids important for NspS function. We performed random mutagenesis of thenspSgene, identified mutant clones deficient in biofilm formation, determined their responsiveness to norspermidine and mapped the location of these residues onto NspS homology models. Single mutants clustered on two lobes of the NspS model, but the majority were found on a single lobe that appeared to be more mobile upon norspermidine binding. We also identified residues in the putative ligand-binding site that may be important for norspermidine binding and interactions with MbaA. Ultimately, our results provide new insights into this novel signalling pathway inV. choleraeand highlight differences between periplasmic binding proteins involved in transport versus signal transduction.