Bypass suppression analysis maps the signalling pathway within a multidomain protein: the RsbP energy stress phosphatase 2C from Bacillus subtilis
Bypass suppression analysis maps the signalling pathway within a multidomain protein: the RsbP energy stress phosphatase 2C from Bacillus subtilis
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DOI：
10.1111/j.1365-2958.2009.06722.x
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发表时间：
2009-06-01
影响因子：
3.6
通讯作者：
Price, Chester W.
中科院分区：
文献类型：
--
作者：
Brody, Margaret S.;Stewart, Valley;Price, Chester W.
枯草芽孢杆菌中控制一般应激反应的网络需要 RsbP 磷酸酶和 RsbQ α/β 水解酶来传递能量应激信号。 RsbP 包含三个结构域：N 端 PAS、中央卷曲螺旋和 C 端 PP2C 磷酸酶。我们在此报告了一项遗传分析，该分析确定了这些结构域的功能相互作用及其与 RsbQ 的关系。 rsbP 的随机诱变产生了 17 个独立的旁路抑制子，它们在 rsbQ 无效菌株背景中具有活性。改变的残基聚集在 RsbP 的三个区域：卷曲螺旋和磷酸酶结构域的两个预测螺旋。一个螺旋 (α 0) 是具有 N 末端传感结构域的细菌 PP2C 磷酸酶亚家族所独有的。另一个 (alpha 1) 与所有已解析的 PP2C 结构中的活性位点不同。抑制子和定向缺失的表型支持这样一种模型，其中卷曲螺旋可能通过 α0-α1 螺旋负向控制磷酸酶活性，其中 RsbQ 水解酶活性和 PAS 结构域共同构成对抗卷曲螺旋的正传感模块。我们认为 α0 螺旋是许多细菌信号蛋白中延伸的 PP2C 结构域的特征，并表明它提供了一种从不同输入结构域传递信息的方法。
The network controlling the general stress response in Bacillus subtilis requires both the RsbP phosphatase and the RsbQ alpha/beta hydrolase to convey signals of energy stress. RsbP contains three domains: an N-terminal PAS, a central coiled-coil and a C-terminal PP2C phosphatase. We report here a genetic analysis that established the functional interactions of the domains and their relationship to RsbQ. Random mutagenesis of rsbP yielded 17 independent bypass suppressors that had activity in an rsbQ null strain background. The altered residues clustered in three regions of RsbP: the coiled-coil and two predicted helices of the phosphatase domain. One helix (alpha 0) is unique to a subfamily of bacterial PP2C phosphatases that possess N-terminal sensing domains. The other (alpha 1) is distinct from the active site in all solved PP2C structures. The phenotypes of the suppressors and directed deletions support a model in which the coiled-coil negatively controls phosphatase activity, perhaps via the alpha 0-alpha 1 helices, with RsbQ hydrolase activity and the PAS domain jointly comprising a positive sensing module that counters the coiled-coil. We propose that the alpha 0 helix characterizes an extended PP2C domain in many bacterial signalling proteins, and suggest it provides a means to communicate information from diverse input domains.