PCR mutagenesis identifies a polymerase-binding sequence of sigma 54 that includes a sigma 70 homology region.
Sigma 54 is a minor bacterial sigma factor that is not a member of the sigma 70 family of proteins but binds the same core RNA polymerase. Previously, we identified a region of sigma 54 that is important for binding core polymerase. In this work, PCR mutagenesis was used to identify specific amino acids important for this binding. The results show that important residues are clustered most closely in a short sequence that was previously speculated to be potentially homologous to a sequence in sigma 70. The mutagenesis also identifies important residues in the flanking hydrophobic-acidic region of sigma 54, which is absent in sigma 70. Overall, the data indicate that sigma 54 binds core polymerase through a sequence homologous to that of sigma 70 but in addition uses unique motifs to modify this interaction.
Tintut
Y
Y
Department of Chemistry and Biochemistry, University of California, Los Angeles 90095, USA.
Gralla
J D
JD
D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015723:Gene Library; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016133:Polymerase Chain Reaction / Q000187:drug effects; D051877:RNA Polymerase Sigma 54; D017386:Sequence Homology, Amino Acid; D012808:Sigma Factor / Q000235:genetics* / Q000378:metabolism*
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D012808:Sigma Factor; C497751:rpoN protein, E coli; D008345:Manganese; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases; D051877:RNA Polymerase Sigma 54; D008274:Magnesium