Oriented DNA binding by one-armed lambda repressor heterodimers and contacts between repressor and RNA polymerase at P(RM).
Bacteriophage lambda repressor activates transcription from P(RM) by contacting the sigma subunit of E. coli RNA polymerase. Although mutations in repressors that are defective in activation affect exposed residues in the repressor-operator co-crystal, the subunit in repressor dimers that is responsible for activation has not been determined experimentally. Here, we describe an oriented heterodimer approach using one-armed repressor-leucine zipper fusion proteins to resolve this question. Protection against methylation of operator DNA in vitro by one-armed and two-armed repressor-leucine zipper heterodimers shows that the subunit containing the arm in a one-armed repressor heterodimer binds preferentially in the consensus half-site of lambda operators. The patterns of activation of the P(RM) promoter in vivo by one-armed heterodimers containing the pc2 mutation in only one subunit show that RNA polymerase at P(RM) contacts the N-terminal domain of the promoter-proximal subunit of the repressor dimer.
Kim
Y I
YI
Department of Biochemistry and Biophysics, Texas A&M University, College Station 77843-2128, USA.
Hu
J C
JC
D010582:Bacteriophage lambda / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D019175:DNA Methylation; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D012097:Repressor Proteins / Q000378:metabolism*; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D012097:Repressor Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D012321:DNA-Directed RNA Polymerases