The C-terminal domain of the novel essential protein Gcp is critical for interaction with another essential protein YeaZ of Staphylococcus aureus.
The C-terminal domain of the novel essential protein Gcp is critical for interaction with another essential protein YeaZ of Staphylococcus aureus.
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DOI：
10.1371/journal.pone.0020163
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发表时间：
2011
期刊：
影响因子：
3.7
通讯作者：
Ji Y
中科院分区：
文献类型：
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作者：
Lei T;Liang X;Yang J;Yan M;Zheng L;Walcheck B;Ji Y
以前的研究已经证明，新的蛋白质Gcp是各种细菌物种（包括金黄色葡萄球菌）生存所必需的;然而，细菌生长所需的原因尚不清楚。为了探讨这种重要性的潜在机制，我们进行了RT-PCR分析，发现gcp基因（sa 1854）与sa 1855，yeaZ（sa 1856）和sa 1857基因共转录，表明这些基因位于同一个操纵子。此外，我们证明了Gcp相互作用与YeaZ使用酵母双杂交（Y2 H）系统和体外下拉测定。为了表征Gcp-YeaZ相互作用，我们对Gcp的C-末端片段的残基进行丙氨酸扫描突变。我们发现C端Y317-F322区域的突变消除了Gcp与YeaZ的相互作用，D324-N329和S332-Y336区域的突变减轻了Gcp与YeaZ的结合。更重要的是，我们证明了Gcp的这些关键区域也是细菌存活所必需的，因为这些突变的Gcp不能补充内源性Gcp的消耗。综上所述，我们的数据表明，Gcp和YeaZ的相互作用可能有助于Gcp对S.金黄色葡萄球菌存活率。我们的发现为这种新型必需蛋白的潜在机制和生物学功能提供了新的见解。
Previous studies have demonstrated that the novel protein Gcp is essential for the viability of various bacterial species including Staphylococcus aureus; however, the reason why it is required for bacterial growth remains unclear. In order to explore the potential mechanisms of this essentiality, we performed RT-PCR analysis and revealed that the gcp gene (sa1854) was co-transcribed with sa1855, yeaZ (sa1856) and sa1857 genes, indicating these genes are located in the same operon. Furthermore, we demonstrated that Gcp interacts with YeaZ using a yeast two-hybrid (Y2H) system and in vitro pull down assays. To characterize the Gcp-YeaZ interaction, we performed alanine scanning mutagenesis on the residues of C-terminal segment of Gcp. We found that the mutations of the C-terminal Y317-F322 region abolished the interaction of Gcp and YeaZ, and the mutations of the D324-N329 and S332-Y336 regions alleviated Gcp binding to YeaZ. More importantly, we demonstrated that these key regions of Gcp are also necessary for the bacterial survival since these mutated Gcp could not complement the depletion of endogenous Gcp. Taken together, our data suggest that the interaction of Gcp and YeaZ may contribute to the essentiality of Gcp for S. aureus survival. Our findings provide new insights into the potential mechanisms and biological functions of this novel essential protein.
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