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  • Title: Structure and properties of a re-engineered homeodomain protein-DNA interface.
    Author: Simon MD, Feldman ME, Rauh D, Maris AE, Wemmer DE, Shokat KM.
    Journal: ACS Chem Biol; 2006 Dec 15; 1(12):755-60. PubMed ID: 17240973.
    Abstract:
    The homeodomain (HD)-DNA interface has been conserved over 500 million years of evolution. Despite this conservation, we have successfully re-engineered the engrailed HD to specifically recognize an unnatural nucleotide using a phage display selection. Here we report the synthesis of novel nucleosides and the selection of mutant HDs that bind these nucleotides using phage display. The high-resolution crystal structure of one mutant in complex with modified and unmodified DNA demonstrates that, even with the substantial perturbation to the interface, this selected mutant retains a canonical HD structure. Dissection of the contributions due to each of the selected mutations reveals that the majority of the modification-specific binding is accomplished by a single mutation (I47G) but that the remaining mutations retune the stability of the HD. These results afford a detailed look at a re-engineered protein-DNA interaction and provide insight into the opportunities for re-engineering highly conserved interfaces.
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