Role of the glutamic and aspartic residues in Na+-ATPase function in the ZrENA1 gene of Zygosaccharomyces rouxii
2002
Watanabe, Y. | Shimono, Y. | Tsuji, H. | Tamai, Y.
The effect of replacement of negatively charged amino acid residues on the function of Na(+) transport proteins of the salt-tolerant yeast Zygosaccharomyces rouxii was examined by heterologous expression of mutant proteins in a strain of Saccharomyces cerevisiae, RH16.6, that lacks native Na(+)-ATPase activity due to null mutations of ENA1, ENA2, ENA3, and ENA4. Mutants of Na(+)/H(+) antiporter gene (ZrSOD2) and Na(+)-ATPase gene (ZrENA1) of Z. rouxii were generated by site-directed mutagenesis. The significance of two aspartic residues arranged in tandem (D265 and D266) was demonstrated in Z. rouxii Na(+)/H(+) antiporter. Some Z rouxii Na(+)-ATPase mutant genes, namely E778A, D852A, and E981A present in the transmembrane domains (TMDs) and D736A, D743A, D748A, D749A, D759A, and D760A present in the cytoplasmic space were constructed. A lower level of salt tolerance was bestowed by the mutant genes D852A and E981A present in TMDs and D748A and D749A present in cytoplasmic space, compared with the wild-type gene (ZrENA1).
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