Characterization of Two Na⁺(K⁺, Li⁺)/H⁺ Antiporters from Natronorubrum daqingense

The Na⁺/H⁺ antiporter NhaC family protein is a kind of Na⁺/H⁺ exchanger from the ion transporter (IT) superfamily, which has mainly been identified in the halophilic bacteria of Bacillus. However, little is known about the Na⁺/H⁺ antiporter NhaC family of proteins in the extremely halophilic archaea. In this study, two Na⁺/H⁺ antiporter genes, nhaC1 and nhaC2, were screened from the genome of Natronorubrum daqingense based on the gene library and complementation of salt-sensitive Escherichia coli KNabc. A clone vector pUC18 containing nhaC1 or nhaC2 could make KNabc tolerate 0.6 M/0.7 M NaCl or 30 mM/40 mM LiCl and a pH of up to 8.5/9.5, respectively. Functional analysis shows that the Na⁺(K⁺, Li⁺)/H⁺ antiport activities of NhaC1 and NhaC2 are both pH-dependent in the range of pH 7.0-10.0, and the optimal pH is 9.5. Phylogenetic analysis shows that both NhaC1 and NhaC2 belong to the Na⁺/H⁺ antiporter NhaC family of proteins and are significantly distant from the identified NhaC proteins from Bacillus. In summary, we have identified two Na⁺(K⁺, Li⁺)/H⁺ antiporters from N. daqingense.