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Related Experiment Videos

Artificial nucleosome positioning sequences.

T E Shrader1, D M Crothers

  • 1Department of Chemistry and Molecular Biophysics, Yale University, New Haven, CT 06511.

Proceedings of the National Academy of Sciences of the United States of America
|October 1, 1989
PubMed
Summary
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Researchers designed DNA sequences that strongly bind to nucleosomes, improving DNA packaging. These novel sequences enhance histone octamer binding and nucleosome formation compared to natural DNA sequences.

Area of Science:

  • Molecular Biology
  • Biophysics
  • Genetics

Background:

  • Nucleosomes are fundamental units of DNA packaging in eukaryotes.
  • DNA sequence significantly influences nucleosome formation and stability.
  • Understanding DNA bendability rules is key to designing functional DNA molecules.

Purpose of the Study:

  • To design and test novel DNA molecules with enhanced nucleosome incorporation.
  • To investigate the sequence dependence of DNA bendability for nucleosome binding.
  • To compare the efficacy of designed sequences against natural nucleosome positioning sequences.

Main Methods:

  • Utilized emerging rules for sequence-dependent DNA bendability.
  • Employed competitive reconstitution experiments for histone octamer binding assays.

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  • Synthesized and tested DNA molecules with repetitive (A/T)3NN(G/C)3NN motifs.
  • Studied the impact of varying lengths of repetitive DNA on nucleosome formation.

    • Main Results:

    • Designed DNA molecules with repeated (A/T)3NN(G/C)3NN motifs showed superior histone octamer binding and nucleosome formation.
    • A 40 bp repetitive DNA segment within a 160 bp fragment was sufficient for strong nucleosome binding.
    • Designed sequences incorporated into nucleosomes up to 100-fold more strongly than bulk DNA.
    • Differential bending free energies were minimal, suggesting weak sequence dependence for DNA distortion energy in nucleosomes.

    Conclusions:

    • Novel DNA sequence motifs can be designed for significantly enhanced nucleosome incorporation.
    • Nucleosome binding is optimized by specific repetitive DNA structures, not just natural sequences.
    • DNA bending energy in nucleosomes is largely independent of sequence, contrary to some expectations.