Sequence analysis and functional studies of a chromosomal region of alkaliphilic Bacillus firmus OF4 encoding an ABC-type transporter with similarity of sequence and Na+ exclusion capacity to the Bacillus subtilis NatAB transporter.
A 14.1-kb DNA fragment was cloned from a lambda library containing inserts of DNA from alkaliphilic Bacillus firmus OF4 on the basis of its hybridization to a probe from a previously sequenced alkaliphile homolog of the natA gene from Bacillus subtilis. Sequence analysis of the entire fragment revealed that, as in B. subtilis, the natA gene was part of a putative gene locus encoding an ABC-type transporter. In the alkaliphile, the transporter involved three genes, designated natCAB, that are part of a larger operon of unknown function. This is in contrast to the two-gene natAB operon and to another homolog from B. subtilis, the yhaQP genes. Like natAB, however, the alkaliphile natCAB catalyzes Na+ extrusion as assessed in a mutant of Escherichia coli that is deficient in Na+ extrusion. The full 14.1-kb fragment of alkaliphile DNA sequenced in this study contained several probable operons as well as likely monocistronic units. Among the 17 predicted ORFs apart from natCAB were acsA, a homolog of a halobacterial gene encoding acetylCoA synthetase; sspA, a homolog of a small acid-soluble spore protein; and malK, an ATP-binding component that was unaccompanied by candidates for other mal transport genes but was able to complement a malK-deficient mutant of E. coli. No strong candidates for genes encoding a secondary Na+/H+ antiporter were found in the fragment, either from the sequence analysis or from analyses of complementation of E. coli mutants by subclones of the 14.1-kb piece. There were a total of 12 ORFs whose closest and significant homologs were genes from B. subtilis; of these, one-third were in apparently different contexts, as assessed by the sequence of the neighboring genes, than the B. subtilis homologs.
Wei
Y
Y
Department of Biochemistry, Mount Sinai School of Medicine, New York, NY 10029, USA.
Guffanti
A A
AA
Krulwich
T A
TA
D018528:ATP-Binding Cassette Transporters / Q000235:genetics*; D026921:Amino Acid Transport Systems, Neutral; D001407:Bacillus / Q000235:genetics*; D001412:Bacillus subtilis / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics; D002876:Chromosomes, Bacterial / Q000502:physiology*; D004269:DNA, Bacterial / Q000032:analysis*; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005816:Genetic Complementation Test; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism; D016296:Mutagenesis; D017422:Sequence Analysis, DNA; D012964:Sodium / Q000378:metabolism*
D018528:ATP-Binding Cassette Transporters; D026921:Amino Acid Transport Systems, Neutral; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; C068021:MalK protein, Bacteria; C489398:MalK protein, E coli; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D012964:Sodium