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  • Title: Isolation of lambda repressor mutants with defects in cooperative operator binding.
    Author: Beckett D, Burz DS, Ackers GK, Sauer RT.
    Journal: Biochemistry; 1993 Sep 07; 32(35):9073-9. PubMed ID: 8369279.
    Abstract:
    A hybrid operator-promoter region was designed to aid in a screen for cooperativity mutants of the lambda repressor. In this system, lambda repressor mutants with defects in pairwise cooperative binding are unable to act as efficient transcriptional repressors. Four single amino acid substitutions in the C-terminal domain of the repressor were isolated. Studies of the DNA binding properties of the purified mutant proteins show that a repressor bearing the Gly147-->Asp mutation binds with normal affinity to single operator sites but is defective in pairwise cooperative site binding. Quantitative footprinting studies show that the free energy of interaction between repressor dimers bound at operator sites OR1 and OR2 is reduced from -2.4 kcal/mol for the wild-type repressor to 0 kcal/mol for the GD147 mutant.
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