Cloning, sequencing and expression of the uvrA gene from an extremely thermophilic bacterium, Thermus thermophilus HB8.
One of the most important DNA repair systems is the nucleotide (nt) excision repair system. The uvr A gene, which plays an essential role in the prokaryotic excision repair system, was cloned from an extremely thermophilic eubacterium, Thermus thermophilus (Tt) HB8, and its nt sequence was determined. In the amino acid (aa) sequence of Tt UvrA, a characteristic duplicated structure, two nt-binding consensus sequences (Walker's A-type motif) and two zinc finger DNA-binding motifs were found. The aa sequence showed 73% homology with that of Escherichia coli (Ec). These features suggest that Tt has the same excision repair system as Ec. Upon comparison of the Tt and Ec UvrA, some characteristic aa substitutions were found. The numbers of Arg and Pro residues were increased (from 66 to 81 and from 47 to 55, respectively), and the numbers of Asn and Met residues were decreased (from 33 to 18 and from 18 to 11, respectively) in Tt. The Tt uvr A gene was expressed in Ec under control of the lac promoter. Purified UvrA was stable up to 80 degrees C (at neutral pH) and at pH 2-11 (at 25 degrees C).
Yamamoto
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N
Department of Biology, Faculty of Science, Osaka University, Japan.
Kato
R
R
Kuramitsu
S
S
D000251:Adenosine Triphosphatases / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D001482:Base Composition; D001483:Base Sequence; D003001:Cloning, Molecular; D004260:DNA Repair / Q000502:physiology; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial / Q000235:genetics*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D016963:Thermus thermophilus / Q000737:chemistry / Q000235:genetics*; D016335:Zinc Fingers
D000596:Amino Acids; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; C068631:UvrA protein, E coli; D000251:Adenosine Triphosphatases