Characterization of a gene responsible for the Na+/H+ antiporter system of alkalophilic Bacillus species strain C-125.
An alkali-sensitive mutant, 38154, of the alkalophilic Bacillus sp. strain C-125 could not grow at an alkaline pH. The nucleotide sequence of a 3.7 kb parental DNA fragment that recovers the growth of 38154 at alkaline pH has four open reading frames (ORF1-4). By subcloning the fragment, we demonstrated that a 0.25 kb DNA region is responsible for the recovery. Direct sequencing of the mutant's corresponding region revealed a G to A substitution. The mutation resulted in an amino acid substitution from Gly-393 to Arg of the putative ORF1 product, which was deduced to be an 804-amino-acid polypeptide with a molecular weight of 89,070. The N-terminal part of the putative ORF1 product showed amino acid similarity to those of the chain-5 products of eukaryotic NADH quinone oxidoreductases. Membrane vesicles prepared from 38154 did not show membrane potential (delta psi)-driven Na+/H+ antiporter activity. Antiporter activity was resumed by introducing a parental DNA fragment which recovered the mutant's alkalophily. These results indicate that the mutation in 38154 affects, either directly or indirectly, the electrogenic Na+/H+ antiporter activity. This is the first report which shows that a gene responsible for the Na+/H+ antiporter system is important in the alkalophily of alkalophilic microorganisms.
Hamamoto
T
T
Institute of Physical and Chemical Research (RIKEN), Saitama, Japan.
Hashimoto
M
M
Hino
M
M
Kitada
M
M
Seto
Y
Y
Kudo
T
T
Horikoshi
K
K
D000595:Amino Acid Sequence; D001407:Bacillus / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport; D008564:Membrane Potentials; D008969:Molecular Sequence Data; D009245:NADH Dehydrogenase; D016366:Open Reading Frames; D010957:Plasmids / Q000235:genetics; D017354:Point Mutation; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D017923:Sodium-Hydrogen Exchangers; C081468:NdhF protein, bacteria; D009245:NADH Dehydrogenase