Characterization of a gene responsible for the Na+/H+ antiporter system of alkalophilic Bacillus species strain C‐125

Abstract

Summary: 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 sub‐cloning 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 0RF1 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 quinine oxidoreductases. Membrane vesicles prepared from 38154 did not show membrane potential (δψ)‐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⁺ anti‐porter system is important in the alkalophily of alkalo‐philic microorganisms.