Identification and Characterization of the Na+/H+ Antiporter NhaS3 from the Thylakoid Membrane of Synechocystis sp. PCC 6803*
Identification and Characterization of the Na+/H+ Antiporter NhaS3 from the Thylakoid Membrane of Synechocystis sp. PCC 6803*
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DOI：
10.1074/jbc.m109.001875
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发表时间：
2009-04
期刊：
影响因子：
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通讯作者：
K. Tsunekawa;T. Shijuku;Mitsuo Hayashimoto;Y. Kojima;K. Onai;M. Morishita;M. Ishiura;T. Kuroda;Tatsunosuke Nakamura;Hiroshi Kobayashi;Mayuko Sato;K. Toyooka;K. Matsuoka;T. Omata;N. Uozumi
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作者：
K. Tsunekawa;T. Shijuku;Mitsuo Hayashimoto;Y. Kojima;K. Onai;M. Morishita;M. Ishiura;T. Kuroda;Tatsunosuke Nakamura;Hiroshi Kobayashi;Mayuko Sato;K. Toyooka;K. Matsuoka;T. Omata;N. Uozumi
Na+/H+ 反向转运蛋白影响质子或钠跨膜动力。集胞藻属。 PCC 6803 有 6 个编码 Na+/H+ 逆向转运蛋白的基因：nhaS1–5 和 sll0556。在本研究中，检测了 NhaS3 的功能。 NhaS3 对于集胞藻的生长至关重要，并且 nhaS3 的缺失不能通过大肠杆菌 Na+/H+ 逆向转运蛋白 NhaA 的表达来补充。膜分级分离、免疫印迹以及免疫金标记显示 NhaS3 位于集胞藻的类囊体膜中。当在大肠杆菌中表达时，NhaS3 在 pH 6.5 至 9.0 的 pH 范围内发挥作用。集胞藻基因组中 nhaS3 拷贝数的减少使得细胞对高 Na+ 浓度更加敏感。 NhaS3 本身没有 K+/H+ 交换活性，但当在大肠杆菌钾吸收突变体中表达时，增强了从培养基中吸收 K+。从低 CO2 条件转变为高 CO2 条件后，nhaS3 的表达增加。 nhaS3 的表达也被发现受昼夜节律控制。基因表达在主观夜晚开始时达到顶峰。这与因释放氧气的光合作用停止而导致二氧化碳消耗率最低的时间一致。这是首次报道位于类囊体膜上的 Na+/H+ 逆向转运蛋白。我们的结果表明，NhaS3 在维持 H+、Na+ 和 K+ 离子稳态中的作用，支持光合产物的转化和黑暗中的能量供应。
Na+/H+ antiporters influence proton or sodium motive force across the membrane. Synechocystis sp. PCC 6803 has six genes encoding Na+/H+ antiporters, nhaS1–5 and sll0556. In this study, the function of NhaS3 was examined. NhaS3 was essential for growth of Synechocystis, and loss of nhaS3 was not complemented by expression of the Escherichia coli Na+/H+ antiporter NhaA. Membrane fractionation followed by immunoblotting as well as immunogold labeling revealed that NhaS3 was localized in the thylakoid membrane of Synechocystis. NhaS3 was shown to be functional over a pH range from pH 6.5 to 9.0 when expressed in E. coli. A reduction in the copy number of nhaS3 in the Synechocystis genome rendered the cells more sensitive to high Na+ concentrations. NhaS3 had no K+/H+ exchange activity itself but enhanced K+ uptake from the medium when expressed in an E. coli potassium uptake mutant. Expression of nhaS3 increased after shifting from low CO2 to high CO2 conditions. Expression of nhaS3 was also found to be controlled by the circadian rhythm. Gene expression peaked at the beginning of subjective night. This coincided with the time of the lowest rate of CO2 consumption caused by the ceasing of O2-evolving photosynthesis. This is the first report of a Na+/H+ antiporter localized in thylakoid membrane. Our results suggested a role of NhaS3 in the maintenance of ion homeostasis of H+, Na+, and K+ in supporting the conversion of photosynthetic products and in the supply of energy in the dark.