Mutant lambda repressors with increased operator affinities reveal new, specific protein-DNA contacts.
The binding specificities of four mutant lambda cI repressor proteins with increased affinities for operator DNA were examined. Two mutant repressors (Glu34----Lys and Glu83----Lys) have the same specificity of binding as wild-type repressor, whereas two (Gly48----Ser and Gly48----Asn) have new binding specificities. The Gly48----Asn mutant repressor recognizes lambda operators with changes at base pair 3 with a different order of affinity than wild-type repressor, suggesting that the side chain of Asn48 makes additional specific DNA contacts at or near this base pair. When paired with a change that disrupts the specific interaction of the amino-terminal arm of lambda repressor with DNA (Lys4----Gln), one change that increases the affinity of repressor (Gly48----Ser) suppresses the binding defect of the Lys4----Gln repressor, resulting in a double mutant repressor with a new binding specificity different than that of both its parents and of wild type. These results lend strong support to the model of direct recognition of the lambda operator by lambda repressor proposed from the crystal structure of the repressor/operator complex.
Benson
N
N
Department of Biological Sciences, University of Southern California, Los Angeles 90089-1481.
Adams
C
C
Youderian
P
P
D010582:Bacteriophage lambda / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites / Q000502:physiology; D016384:Consensus Sequence; D004279:DNA, Viral / Q000378:metabolism*; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics; D008242:Lysogeny / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009875:Operator Regions, Genetic / Q000502:physiology*; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins