Isolation of the rpoD gene encoding the principal sigma factor of the deep-sea piezophilic bacterium Shewanella violacea strain DSS12 and its overexpression in Escherichia coli.
The gene encoding the principal a factor (rpoD) of the piezophilic bacterium Shewanella violacea was cloned and sequenced. The rpoD gene was found to encode a polypeptide consisting of 614 amino acid residues, showing 75.6 and 64.3% identity to those of Escherichia coli and Pseudomonas putida, respectively. Comparison with E. coli sigma70 and P. putida sigma70 showed that significant similarity exists in four conserved regions known to be required for promoter recognition and core binding. Using an expression plasmid harboring the rpoD gene, the S. violacea sigma70 factor was overexpressed in E. coli and successfully purified to near homogeneity.
The DEEP STAR Group, Japan Marine Science and Technology Center, Kanagawa, Japan. nakasone@jamstec.go.jp
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Nakasone
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Ikegami
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Kato
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Horikoshi
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D020592:Shewanella / Q000235:genetics*; D012808:Sigma Factor / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012808:Sigma Factor; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases