Journal
JOURNAL OF MOLECULAR BIOLOGY
Volume 425, Issue 1, Pages 133-143Publisher
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2012.10.008
Keywords
DNA binding; DNA structure; indirect readout; bacteriophage
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Funding
- College of Arts and Sciences at the University at Buffalo
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The repressor of bacteriophage P22 (P22R) discriminates between its various DNA binding sites by sensing the identity of non-contacted base pairs at the center of its binding site. The indirect readout of these non-contacted bases is apparently based on DNA's sequence-dependent conformational preferences. The structures of P22R-DNA complexes indicate that the non-contacted base pairs at the center of the binding site are in the B' state. This finding suggests that indirect readout and therefore binding site discrimination depend on P22R's ability to either sense and/or impose the B' state on the non-contacted bases of its binding sites. We show here that the affinity of binding sites for P22R depends on the tendency of the central bases to assume the B'-DNA state. Furthermore, we identify functional groups in the minor groove of the non-contacted bases as the essential modulators of indirect readout by P22R. In P22R DNA complexes, the negatively charged E44 and E48 residues are provocatively positioned near the negatively charged DNA phosphates of the non-contacted nucleotides. The close proximity of the negatively charged groups on protein and DNA suggests that electrostatics may play a key role in the indirect readout process. Changing either of two negatively charged residues to uncharged residues eliminates the ability of P22R to impose structural changes on DNA and to recognize non-contacted base sequence. These findings suggest that these negatively charged amino acids function to force the P22R-bound DNA into the B' state and therefore play a key role in indirect readout by P22R. (C) 2012 Elsevier Ltd. All rights reserved.
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