P22 Arc Repressor: Role of Cooperativity in Repression and Binding to Operators with Altered Half-site Spacing

Abstract

Dimers of P22 Arc repressor bind to half-sites of the 21 bparcoperator and interact cooperatively to stabilize a DNA-bound tetramer. Mutation of Ser35 (a residue in the dimer–dimer interface) to Arg or Leu disrupts cooperative binding. The mutant proteins have near wild-type stabilities, give operator footprints like wild-type, and prevent binding of RNA polymerase to the Pantpromoterin vitro. These mutants are, however, largely inactivein vivo. Thus, although cooperativity is not structurally required for repression, it appears that the additional DNA-binding energy from dimer–dimer cooperativity is required for normal biological function. Altering the spacing between the DNA half-sites by even one base-pair eliminates dimer–dimer cooperativity, indicating that Arc dimers need to be oriented correctly by half-site binding to allow the interactions that stabilize the tetrameric complex.