Genetic identification of three ABC transporters as essential elements for nitrate respiration in Haloferax volcanii.
More than 40 nitrate respiration-deficient mutants of Haloferax volcanii belonging to three different phenotypic classes were isolated. All 15 mutants of the null phenotype were complemented with a genomic library of the wild type. Wild-type copies of mutated genes were recovered from complemented mutants using two different approaches. The DNA sequences of 13 isolated fragments were determined. Five fragments were found to overlap; therefore nine different genomic regions containing genes essential for nitrate respiration could be identified. Three genomic regions containing genes coding for subunits of ABC transporters were further characterized. In two cases, genes coding for an ATP-binding subunit and a permease subunit were clustered and overlapped by four nucleotides. The third gene for a permease subunit had no additional ABC transporter gene in proximity. One ABC transporter was found to be glucose specific. The mutant reveals that the ABC transporter solely mediates anaerobic glucose transport. Based on sequence similarity, the second ABC transporter is proposed to be molybdate specific, explaining its essential role in nitrate respiration. The third ABC transporter is proposed to be anion specific. Genome sequencing has shown that ABC transporters are widespread in Archaea. Nevertheless, this study represents only the second example of a functional characterization.
Wanner
C
C
Biozentrum Niederursel, Institut für Mikrobiologie, J. W. Goethe-Universität D-60439 Frankfurt, Germany.
Soppa
J
J
D018528:ATP-Binding Cassette Transporters / Q000235:genetics* / Q000502:physiology; D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000838:Anions / Q000378:metabolism; D019843:Archaeal Proteins / Q000235:genetics* / Q000502:physiology; D001120:Arginine / Q000032:analysis; D001665:Binding Sites; D004734:Energy Metabolism / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005816:Genetic Complementation Test; D005947:Glucose / Q000378:metabolism; D019622:Haloferax volcanii / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000378:metabolism; D009566:Nitrates / Q000378:metabolism*; D016366:Open Reading Frames; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
D018528:ATP-Binding Cassette Transporters; D000838:Anions; D019843:Archaeal Proteins; D009566:Nitrates; C044659:molybdate; D008982:Molybdenum; D000255:Adenosine Triphosphate; D001120:Arginine; D005947:Glucose