Characterization of a novel two-component Na(+)(Li(+), K(+))/H(+) antiporter from Halomonas zhaodongensis.
Characterization of a novel two-component Na(+)(Li(+), K(+))/H(+) antiporter from Halomonas zhaodongensis.
复制标题
赵东盐单胞菌新型双组分 Na (Li ,K )/H 逆向转运蛋白的表征
DOI：
10.1038/s41598-017-04236-0
复制
发表时间：
2017-06-26
影响因子：
4.6
通讯作者：
Jiang J
中科院分区：
文献类型：
--
作者：
Meng L;Meng F;Zhang R;Zhang Z;Dong P;Sun K;Chen J;Zhang W;Yan M;Li J;Abdel-Motaal H;Jiang J
本研究通过在缺乏三种主要Na+/H+逆向转运蛋白的大肠杆菌KNabc中进行选择，从赵东盐单胞菌的基因组DNA中筛选出新型Na+/H+逆向转运蛋白基因。两个名为 umpAB 的基因的共表达，编码属于 DUF1538（No. 1538 功能未知的结构域）家族的配对同源未知膜蛋白，被发现赋予大肠杆菌 KNabc 对 0.4 M NaCl 和 30 mM LiCl 的耐受性，以及 8.0 的碱性 pH 耐受性。蛋白质印迹和免疫共沉淀证实 UmpAB 以异二聚体形式定位在细胞质膜中。功能分析表明，UmpAB 在 6.5 至 9.5 的广泛 pH 范围内表现出 pH 依赖性 Na+(Li+, K+)/H+ 反向转运活性，最佳 pH 值为 9.0。 UmpA 和 UmpB 均未与已知的单基因或多基因 Na+/H+ 逆向转运蛋白或具有 Na+/H+ 逆向转运活性的 ChaA、MdfA、TetA(L)、Nap 和 PsmrAB 等蛋白质表现出同源性。系统发育分析证实UmpAB应该属于DUF1538家族，该家族与上述具有Na+/H+反向转运活性的蛋白有显着的距离。综上所述，我们提出 UmpAB 代表一种新型的双组分 Na+(Li+, K+)/H+ 反向转运蛋白。据我们所知，这是第一份关于 DUF1538 家族未知膜蛋白功能分析的报告。
In this study, genomic DNA was screened for novel Na+/H+ antiporter genes from Halomonas zhaodongensis by selection in Escherichia coli KNabc lacking three major Na+/H+ antiporters. Co-expression of two genes designated umpAB, encoding paired homologous unknown membrane proteins belonging to DUF1538 (domain of unknown function with No. 1538) family, were found to confer E. coli KNabc the tolerance to 0.4 M NaCl and 30 mM LiCl, and an alkaline pH resistance at 8.0. Western blot and co-immunoprecipitation establish that UmpAB localize as a hetero-dimer in the cytoplasmic membranes. Functional analysis reveals that UmpAB exhibit pH-dependent Na+(Li+, K+)/H+ antiport activity at a wide pH range of 6.5 to 9.5 with an optimal pH at 9.0. Neither UmpA nor UmpB showed homology with known single-gene or multi-gene Na+/H+ antiporters, or such proteins as ChaA, MdfA, TetA(L), Nap and PsmrAB with Na+/H+ antiport activity. Phylogenetic analysis confirms that UmpAB should belong to DUF1538 family, which are significantly distant with the above-mentioned proteins with Na+/H+ antiport activity. Taken together, we propose that UmpAB represent a novel two-component Na+(Li+, K+)/H+ antiporter. To the best of our knowledge, this is the first report on the functional analysis of unknown membrane proteins belonging to DUF1538 family.