Determinants for the activation and autoinhibition of the diguanylate cyclase response regulator WspR.
Determinants for the activation and autoinhibition of the diguanylate cyclase response regulator WspR.
复制标题
二甘氨酸酸环化酶反应调节剂WSPR激活和自抑制的决定因素。
DOI：
10.1016/j.jmb.2009.08.030
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发表时间：
2009-10-30
影响因子：
5.6
通讯作者：
Sondermann, Holger
中科院分区：
文献类型：
--
作者：
De, Nabanita;Navarro, Marcos V. A. S.;Raghavan, Rahul V.;Sondermann, Holger
细菌第二信使c-di-GMP控制细胞的分泌、黏附和运动，导致生物膜的形成和细胞毒性的增加。含有GGDEF的双鸟苷环化酶和含有EAL或HD-GYP结构域的磷酸二酯酶被认为是控制细胞c-di-GMP水平的酶，但关于调控和信号特异性的分子机制尚不清楚。我们最近从假单胞菌那里确定了二鸟苷环化酶反应调节因子WSPR的产物抑制途径，这是一种控制生物膜形成的有效分子开关。在WSPR中，催化活性受到连接其磷酸受体(REC)和GGDEF结构域的螺旋茎基序的调节。茎促进形成不同的寡聚体状态，有助于激活和自我抑制。在这里，我们基于全长WSPR的晶体结构、分离的GGDEF结构域和人工二聚的催化结构域，为WSPR中二鸟苷环化酶活性的调节提供了新的见解。这些结构强调了抑制是通过限制刚性GGDEF结构域的迁移率来实现的，这是通过c-di-GMP与GGDEF结构域上的抑制位点结合来实现的。动力学测量和生化表征证实了WSPR的活化需要形成四聚体物种的模型。四聚在高蛋白浓度下自发发生，或在添加仿磷化合物氟化铍时自发发生。我们的分析阐明了常见的和WSPR特有的微调二鸟苷环化酶活性的机制。
The bacterial second messenger c-di-GMP controls secretion, cell adhesion and motility leading to biofilm formation and increased cytotoxicity. Diguanylate cyclases containing GGDEF and phosphodiesterases containing EAL or HD-GYP domains have been identified as the enzymes controlling cellular c-di-GMP levels, yet less is known regarding the molecular mechanisms governing regulation and signaling specificity. We recently determined a product-inhibition pathway for the diguanylate cyclase response regulator WspR from Pseudomonas, a potent molecular switch that controls biofilm formation. In WspR, catalytic activity is modulated by a helical stalk motif that connects its phospho-receiver (REC) and GGDEF domains. The stalks facilitate the formation of distinct oligomeric states that contribute to both activation and autoinhibition. Here, we provide novel insights into the regulation of diguanylate cyclase activity in WspR based on the crystal structures of full-length WspR, the isolated GGDEF domain, and an artificially dimerized catalytic domain. The structures highlight that inhibition is achieved by restricting the mobility of rigid GGDEF domains, mediated by c-di-GMP binding to an inhibitory site at the GGDEF domain. Kinetic measurements and biochemical characterization corroborate a model in which the activation of WspR requires the formation of a tetrameric species. Tetramerization occurs spontaneously at high protein concentration or upon addition of the phosphomimetic compound beryllium fluoride. Our analyses elucidate common and WspR-specific mechanisms for the fine-tuning of diguanylate cyclase activity.
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DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
3.3
作者：
Goymer, Patrick;Kahn, Sophie G.;Rainey, Paul B.
通讯作者：
Rainey, Paul B.
影响因子：
9.8
作者：
De N;Pirruccello M;Krasteva PV;Bae N;Raghavan RV;Sondermann H
通讯作者：
Sondermann H
DOI：
10.1107/s0907444998003254
发表时间：
1998-09-01
期刊：
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY
影响因子：
--
作者：
Brunger, AT;Adams, PD;Warren, GL
通讯作者：
Warren, GL
DOI：
10.1073/pnas.0511090103
发表时间：
2006-02-21
影响因子：
11.1
作者：
Kulesekara, H;Lee, V;Lory, S
通讯作者：
Lory, S