Cloning and identification of a novel NhaD-type Na+/H+ antiporter from metagenomic DNA of the halophilic bacteria in soil samples around Daban Salt Lake
Cloning and identification of a novel NhaD-type Na+/H+ antiporter from metagenomic DNA of the halophilic bacteria in soil samples around Daban Salt Lake
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达坂盐湖周边土壤嗜盐细菌宏基因组DNA中新型NhaD型Na/H逆向转运蛋白的克隆与鉴定
DOI：
10.1007/s00792-013-0600-2
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发表时间：
2013
期刊：
影响因子：
2.9
通讯作者：
Juquan Jiang
中科院分区：
文献类型：
--
作者：
Lifu Yang;Zhenhui Wang;Lei Wang;Ren Mu;Zhi Zou;Kun Yuan;Yuekun Wang;Haiping Wu;Juquan Jiang
本研究通过在缺乏三种主要Na（+）/H（+）反向转运蛋白的大肠杆菌KNabc中筛选，从大板盐湖嗜盐菌的宏基因组DNA中筛选Na（+）/H（+）反向转运蛋白基因。最终克隆了一个编码NhaD型Na（+）/H（+）逆向转运蛋白的基因，命名为Hb_nhaD。Hb_NhaD的存在使E. coli KNabc在0.5M NaCl和0.2M LiCl及碱性pH条件下，Hb_NhaD的同源性最高（70.6%）与梭菌科一个未知的NhaD型Na（+）/H（+）反向转运蛋白同源性较高，与长盐单胞菌（Halomonaselongata）的NhaD型Na（+）/H（+）反向转运蛋白同源性较低20.8%、解淀粉碱单胞菌19.0%、副溶血性弧菌18.9%、霍乱弧菌18.7%。pH依赖性Na（+）（Li（+））/H（+）逆向转运活性从E. coli KNabc中表达。Hb_NhaD在pH6.5 ~ 9.0范围内表现出很高的Na（+）（Li（+））/H（+）反向转运活性，在pH7.0时活性最高，这与上述已知的NhaD型Na（+）/H（+）反向转运蛋白的活性显著不同。在pH 7.0时，Hb_NhaD对Na（+）和Li（+）的表观Km值分别为1.31和2.16。基于上述结果，我们认为Hb_NhaD是一种新型的NhaD型Na（+）/H（+）逆向转运蛋白。
In this study, metagenomic DNA was screened for the Na(+)/H(+) antiporter gene from the halophilic bacteria in Daban Salt Lake by selection in Escherichia coli KNabc lacking three major Na(+)/H(+) antiporters. One gene designated as Hb_nhaD encoding a novel NhaD-type Na(+)/H(+) antiporter was finally cloned. The presence of Hb_NhaD conferred tolerance of E. coli KNabc to up to 0.5 M NaCl and 0.2 M LiCl, and an alkaline pH. Hb_NhaD has the highest identity (70.6%) with a putative NhaD-type Na(+)/H(+) antiporter from an uncharacterized Clostridiaceae species, and also has lower identity with known NhaD-type Na(+)/H(+) antiporters from Halomonas elongata (20.8%), Alkalimonas amylolytica (19.0%), Vibrio parahaemolyticus (18.9%) and Vibrio cholerae (18.7 %). pH-dependent Na(+)(Li(+))/H(+) antiport activity was detected from everted membrane vesicles prepared from E. coli KNabc carrying Hb_nhaD. Hb_NhaD exhibited very high Na(+)(Li(+))/H(+) antiport activity over a wide pH range from 6.5 to 9.0 with the highest activity at pH 7.0 which is significantly different from those of the above known NhaD-type Na(+)/H(+) antiporters. Also, the apparent K m values of Hb_NhaD for Na(+) and Li(+) at pH 7.0 were determined to be 1.31 and 2.16, respectively. Based on the above results, we proposed that Hb_NhaD should be categorized as a novel NhaD-type Na(+)/H(+) antiporter.
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DOI：
10.1186/1746-1448-2-10
发表时间：
2006-07-27
期刊：
Saline systems
影响因子：
--
作者：
Kurz M;Brünig AN;Galinski EA
通讯作者：
Galinski EA
DOI：
10.1016/s0021-9258(18)42779-2
发表时间：
1992-03
期刊：
The Journal of biological chemistry
影响因子：
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作者：
M. Waser;Denise Hess-BienzS;K. Davies;Marc Soliozg
通讯作者：
M. Waser;Denise Hess-BienzS;K. Davies;Marc Soliozg
DOI：
10.1016/s0167-4781(98)00058-x
发表时间：
1998-07
期刊：
Biochimica et biophysica acta
影响因子：
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作者：
J. Utsugi;K. Inaba;T. Kuroda;M. Tsuda;T. Tsuchiya
通讯作者：
J. Utsugi;K. Inaba;T. Kuroda;M. Tsuda;T. Tsuchiya
DOI：
10.1016/0304-4173(83)90011-3
发表时间：
1983-12
期刊：
Biochimica et biophysica acta
影响因子：
--
作者：
T. A. Krulwich
通讯作者：
T. A. Krulwich
DOI：
10.1016/s0021-9258(18)46994-3
发表时间：
1994-11
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
J. Cheng;A. Guffanti;T. A. Krulwich
通讯作者：
J. Cheng;A. Guffanti;T. A. Krulwich