Involvement of WalK (VicK) phosphatase activity in setting WalR (VicR) response regulator phosphorylation level and limiting cross-talk in Streptococcus pneumoniae D39 cells.
Involvement of WalK (VicK) phosphatase activity in setting WalR (VicR) response regulator phosphorylation level and limiting cross-talk in Streptococcus pneumoniae D39 cells.
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DOI：
10.1111/mmi.12006
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发表时间：
2012-11
影响因子：
3.6
通讯作者：
Winkler ME
中科院分区：
文献类型：
--
作者：
Wayne KJ;Li S;Kazmierczak KM;Tsui HC;Winkler ME
低GC革兰氏阳性菌的WalRK（YycFG）双组分系统（TCS）在调节参与细胞分裂和壁应激反应的肽聚糖水解酶基因中起关键作用。肺炎链球菌（肺炎球菌）和其他链球菌菌种的WalRK（VicRK）TCS与其他低GC菌种的TCS存在许多差异。值得注意的是，肺炎球菌WalK传感器激酶对于培养物中的正常生长不是必需的，不像其在芽孢杆菌属和葡萄球菌属物种中的同源物。WalK传感器激酶具有组氨酸自激酶活性，并介导磷酸化WalR~P反应调节剂的去磷酸化。为了了解这两个WalK活性对肺炎球菌生长的贡献，我们构建并表征了一组在生化反应和细胞中的WalK激酶和磷酸酶突变体。我们在WalK中发现了一个氨基酸取代，它显著降低了磷酸酶活性，但没有降低其他活性。通过Phos-tag SDS-PAGE测定细胞中WalRK调节子表达水平和WalR~P量之间的比较。WalK磷酸酶活性的降低导致近90%的磷酸化为WalR~P，这与WalK磷酸酶在指数生长的细胞中具有强烈活性的结论一致。WalK磷酸酶活性也被证明依赖于WalK PAS结构域并限制串扰和WalK+细胞中WalR~P的恢复。
WalRK (YycFG) two-component systems (TCSs) of low-GC Gram-positive bacteria play critical roles in regulating peptidoglycan hydrolase genes involved in cell division and wall stress responses. The WalRK (VicRK) TCSs of Streptococcus pneumoniae (pneumococcus) and other Streptococcus species show numerous differences with those of other low-GC species. Notably, the pneumococcal WalK sensor kinase is not essential for normal growth in culture, unlike its homologues in Bacillus and Staphylococcus species. The WalK sensor kinase possesses histidine autokinase activity and mediates dephosphorylation of phosphorylated WalR~P response regulator. To understand the contributions of these two WalK activities to pneumococcal growth, we constructed and characterized a set of walK kinase and phosphatase mutants in biochemical reactions and in cells. We identified an amino acid substitution in WalK that significantly reduces phosphatase activity, but not other activities. Comparisons were made between WalRK regulon expression levels and WalR~P amounts in cells determined by Phos-tag SDS-PAGE. Reduction of WalK phosphatase activity resulted in nearly 90% phosphorylation to WalR~P, consistent with the conclusion that WalK phosphatase is strongly active in exponentially growing cells. WalK phosphatase activity was also shown to depend on the WalK PAS domain and to limit crosstalk and the recovery of WalR~P from walK+ cells.
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影响因子：
3.6
作者：
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通讯作者：
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