Strong DNA binding by covalently linked dimeric Lac headpiece: Evidence for the crucial role of the hinge helices

Charalampos G. Kalodimos, Gert E. Folkers, Rolf Boelens, Robert Kaptein

Research output: Contribution to journalArticlepeer-review

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Abstract

The combined structural and biochemical studies on Lac repressor bound to operator DNA have demonstrated the central role of the hinge helices in operator bending and the induction mechanism. We have constructed a covalently linked dimeric Lac-headpiece that binds DNA with four orders of magnitude higher affinity as compared with the monomeric form. This enabled a detailed biochemical and structural study of Lac binding to its cognate wild-type and selected DNA operators. The results indicate a profound contribution of hinge helices to the stability of the protein-DNA complex and highlight their central role in operator recognition. Furthermore, protein-DNA interactions in the minor groove appear to modulate hinge helix stability, thus accounting for affinity differences and protein-induced DNA bending among the various operator sites. Interestingly, the in vitro DNA-binding affinity of the reported dimeric Lac construct can de readily modulated by simple adjustment of redox conditions, thus rendering it a potential artificial gene regulator.

Original languageEnglish (US)
Pages (from-to)6039-6044
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume98
Issue number11
DOIs
StatePublished - May 22 2001

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