DNA twisting and the effects of non-contacted bases on affinity of 434 operator for 434 represser

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Abstract
THE bacteriophage 434 represser regulates gene expression by binding with differing affinities to the six operator sites on the phage chromosome1,2. The symmetrically arrayed outer eight base pairs (four in each half-site) of these 14-base-pair operators are highly conserved but the middle four bases are divergent3. Although these four base pairs are not in contact with represser4,5, operators with A·T or T·A base pairs at these positions bind represser more strongly than those bearing C·G or G·C5, suggestiong that these bases are important for the represser's ability to discriminate between operators. There is evidence that the central base pairs influence operator function by constraining the twisting and/or bending of DNA5–7. Here we show that there is a relationship between the intrinsic twist of an operator, as determined by the sequence of its central bases, and its affinity for represser; an operator with a lower affinity is undertwisted relative to an operator with higher affinity. In complex with repressor, the twist of both high- and low-affinity operators is the same. These results indicate that the intrinsic twist of DNA and its twisting flexibility both affect the affinity of 434 operator for repressor.