PRODUCING POSITIVE, NEGATIVE, AND NO COOPERATIVITY BY MUTATIONS AT A SINGLE RESIDUE LOCATED AT THE SUBUNIT INTERFACE IN THE ASPARTATE RECEPTOR OF SALMONELLA-TYPHIMURIUM

Site-directed mutagenesis of the aspartate receptor of Salmonella typh imurium (Tar(s))(1) at serine 68, a residue located within the asparta te binding pocket and at the subunit interface, identified this residu e as an allosteric switch in this receptor. Substitutions at this posi tion can affect both the type and degree of binding cooperativity obse rved. Negative cooperativity is observed in the wild-type receptor (H- n 0.7 +/- 0.1) and is maintained by the mutations S68C (n(H) = 0.8 +/- 0.02), S68V (n(H) = 0.9 +/- 0.05), and S68D (half-of-the-sites). Bind ing at only half of the sites was detectable in the S68D mutant, an ex treme form of negative cooperativity. No cooperativity (n(H) = 1.0 +/- 0.03) was observed in the mutant S68A. Positive cooperativity was gen erated by the substitutions S68T (n(H) = 1.2 +/- 0.09), S68L (n(H) = 1 .2 +/- 0.1), S68N (n(H) = 1.3 +/- 0.2), and S68I (n(H) = 1.4 +/- 0.2). Binding measurements indicated that the substitutions S68Q, S68E, and S68F decrease affinity of the first ligand binding 500-fold, 7000-fol d, and 1600-fold, respectively.