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  • Title: DNA affinity cleaving analysis of homeodomain-DNA interaction: identification of homeodomain consensus sites in genomic DNA.
    Author: Shang Z, Ebright YW, Iler N, Pendergrast PS, Echelard Y, McMahon AP, Ebright RH, Abate C.
    Journal: Proc Natl Acad Sci U S A; 1994 Jan 04; 91(1):118-22. PubMed ID: 7904065.
    Abstract:
    We have incorporated the DNA-cleaving moiety o-phenanthroline-copper at amino acid 10 of the Msx-1 homeodomain, and we have analyzed site-specific DNA cleavage by the resulting Msx-1 derivative. We show that amino acid 10 of the Msx-1 homeodomain is close to the 5' end of the consensus DNA site 5'-(C/G)TAATTG-3' in the Msx-1-DNA complex. Our results indicate that the orientation of the Msx-1 homeodomain relative to DNA is analogous to the orientation of the engrailed and Antennapedia homeodomains. We show further that DNA affinity cleaving permits identification of consensus DNA sites for Msx-1 in kilobase DNA substrates. The specificity of the approach enabled us to identify an Msx-1 consensus DNA site within the transcriptional control region of the developmental regulatory gene Wnt-1. We propose that incorporation of o-phenanthroline-copper at amino acid 10 of a homeodomain may provide a generalizable strategy to determine the orientation of a homeodomain relative to DNA and to identify homeodomain consensus DNA sites in genomic DNA.
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