Sequence specificity of bacteriophage 434 repressor-operator complexation.

Abstract

The binding affinity of the bacteriophage 434 repressor for its DNA operator depends strongly on the nature of two central base-pairs that are not in contact with the dimeric protein. In order to investigate the origin of this sequence specificity, we carried out molecular modelling of five model operators with central TA, AT, CG, GC and IC sequences. The five oligomers were studied both before and after complexation with the N-terminal binding domain of the 434 repressor. The relative importance of nucleic acid flexibility on operator-repressor binding was studied via a low frequency normal mode analysis using an internal coordinate method that we developed recently. The results suggest a higher twisting flexibility for TA and AT central steps than for CG, GC or IC steps, but the differences appear to be too small to account for the strength of repressor binding. An energetic analysis of the model operator-repressor complexes reveals rather that the preference for A.T and T.A base-pairs is electrostatic in origin and is linked to the presence of cationic Arg43 side-chains of repressor. This conclusion is supported by comparison with an R43A mutant and correlates with the sequence dependence of the electrostatic potential in the central minor groove of the operators.