Open AccessExpression of the Na + /H + antiporter gene (g1‐ nhaC ) of alkaliphilic Bacillus sp. G1 in Escherichia coli
Author(s) -
Wai Liew Chong,
Illias Rosli Md.,
Muhammad Mahadi Nor,
Najimudin Nazalan
Publication year - 2007
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2007.00925.x
Subject(s) - antiporter , escherichia coli , recombinant dna , peptide sequence , bacteria , microbiology and biotechnology , biology , chemistry , biochemistry , gene , genetics , membrane
A Na + /H + antiporter gene was isolated from alkaliphilic Bacillus sp. G1. The full‐length sequence of the Na + /H + antiporter gene was obtained using a genome walking method, and designated as g1‐ nhaC . An ORF preceded by a promoter‐like sequence and a Shine–Dalgarno sequence, and followed by a terminator‐like sequence was identified. The deduced amino acid sequence consists of 535 amino acids, and a calculated molecular mass of 57 776 Da. g1‐ nhaC was subsequently cloned into pET22b(+) and expressed in Escherichia coli BL21 (DE3). Recombinant E. coli harboring the g1‐ nhaC gene was able to grow in modified L medium at various concentrations of NaCl (0.2–2.0 M) at different pH values. The recombinant bacteria grew well in the medium with concentrations of NaCl as high as 1.75 M at pH 8.0–9.0. Minimal growth was observed at 2.0 M NaCl, pH 8.0–9.0. At pH 10, the recombinant bacteria grew well in a medium with a low concentration of NaCl (0.2 M). These results suggested that the g1‐NhaC antiporter from Bacillus sp. G1 plays a role in Na + extrusion at lower pH values and in pH homeostasis at pH 10 under Na + ‐limiting conditions.