title: 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.
abstract: 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.
authors.affiliation: Rubber Research Institute, Chinese Academy of Tropical Agricultural Sciences, and Key Laboratory of Rubber Biology and Genetic Resource Utilization, Ministry of Agriculture, Hainan, 571737, Hainan, People's Republic of China.
authors.forename: Hua
authors.initials: H
authors.lastname: Zhang
authors.forename: Zhenhui
authors.initials: Z
authors.lastname: Wang
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.forename: Ren
authors.initials: R
authors.lastname: Mu
authors.forename: Zhi
authors.initials: Z
authors.lastname: Zou
authors.forename: Kun
authors.initials: K
authors.lastname: Yuan
authors.forename: Yuekun
authors.initials: Y
authors.lastname: Wang
authors.forename: Haiping
authors.initials: H
authors.lastname: Wu
authors.forename: Juquan
authors.initials: J
authors.lastname: Jiang
authors.forename: Lifu
authors.initials: L
authors.lastname: Yang
mesh_terms: D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D016680:Genome, Bacterial; D020632:Halomonas / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D054712:Salinity; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics* / Q000378:metabolism; D012988:Soil Microbiology; D014733:Vibrio / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D017923:Sodium-Hydrogen Exchangers