DNA-BASED LOSS OF SPECIFICITY MUTATIONS - EFFECTS OF DNA-SEQUENCE ON THE CONTACTED AND NONCONTACTED BASE PREFERENCES OF BACTERIOPHAGE-P22 REPRESSOR

Although the two central bases of the P22 operator are not contacted b y the P22 repressor, changes in these bases alter the affinity of oper ator for repressor. Previous studies (Wu, L., and Koudelka, G. B. (199 3) J. Biol. Chem. 268, 18975-18981) show that the structure of the P22 repressor-operator complex varies with central base sequence. Here we show that central base sequence composition affects the strength of t wo, and likely all, specific amino acid-base pair contacts between syn thetic P22 operators and P22 repressor. However, alter ing a specific protein-DNA contact via a loss-of-contact mutation in repressor result s in a loss of specificity at only one contacted position. Thus, only changing the sequence of non-contacted bases affects repressor's globa l base specificity. The observed effects of ionic concentration on the affinities of various operators for repressor and the DNase I pattern s of protein complexes with these binding sites indicate certain centr al base sequences facilitate optimal juxtaposition of repressor with i ts contacted bases, while others prevent it. The existence of differen t structural forms of the repressor-operator complexes explains how th e relative energetic importance of specific amino acid-base pair edge contacts is modulated.