Heteronuclear (1H, 13C, 15N) NMR assignments and secondary structure of the basic region-helix-loop-helix domain of E47

Authors

  • Robert Fairman,

    Corresponding author
    1. The DuPont Merck Pharmaceutical Company, Experimental Station, P.O. Box 80500, Wilmington, Delaware 19880–0500
    Current affiliation:
    1. Division of Macromolecular Structure, Bristol-Myers Squibb Pharmaceutical Research Institute, P.O. Box 4000, Princeton, New Jersey 08543–4000
    • Division of Macromolecular Structure, Bristol-Myers Squibb Pharmaceutical Research Institute, P.O. Box 4000, Princeton, New Jersey 08543–4000
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  • Rita K. Beran-Steed,

    1. The DuPont Merck Pharmaceutical Company, Experimental Station, P.O. Box 80500, Wilmington, Delaware 19880–0500
    Current affiliation:
    1. Pharmacia Biotech Inc., P.O. Box 1327, Piscataway, New Jersey 08855–1327
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  • Tracy M. Handel

    Corresponding author
    1. The DuPont Merck Pharmaceutical Company, Experimental Station, P.O. Box 80500, Wilmington, Delaware 19880–0500
    Current affiliation:
    1. Department of Molecular and Cell Biology, University of California, Berkeley, California 94720
    • Department of Molecular and Cell Biology, University of California, Berkeley, Virus Laboratory, 229 Wendell M. Stanley Hall, Berkeley, California 94720
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Abstract

E47 is an immunoglobulin enhancer DNA-binding protein that contains a basic region-helix-loop-helix (b/HLH) domain. This structural motif defines a class of transcription factors that are central to the developmental regulation of many tissues. Its function is to provide a dimerization interface through the formation of a parallel four-helix bundle, resulting in the juxtaposition of two basic DNA-recognition α-helices that control sequence-specific DNA-binding. In order to gain insight into the biophysical nature of b/HLH domains, we have initiated structural studies of the E47 homodimer by NMR. Sequence-specific resonance assignments have been obtained using a combination of heteronuclear double- and triple-resonance NMR experiments. The secondary structure was deduced from characteristic patterns of NOEs, 13Cα/β chemical shifts, and measurements of 3JHNHα scalar couplings. Except for the basic region recognition helix, the secondary structural elements of the E47 homodimer are preserved in the absence DNA when compared with the co-crystal structure of E47 bound to DNA (Ellenberger T, Fass D, Arnaud M, Harrison SC, 1994, Genes & Dev 8:970–980). As expected, the DNA-binding helix is largely unstructured, but does show evidence of nascent helix formation. The HLH region of E47 is structured, but highly dynamic as judged by the rapid exchange of backbone hydrogen atoms and the relatively weak intensities of many of the NOEs defining the dimerization helices. This dynamic nature may be relevant to the ability of E47 both to homodimerize and to heterodimerize with MyoD, Id, and Tal1.

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