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Related Experiment Videos

Predicting coupling limits from an experimentally determined energy landscape.

Timothy O Street1, Christina M Bradley, Doug Barrick

  • 1T. C. Jenkins Department of Biophysics, The Johns Hopkins University, 3400 North Charles Street, Baltimore, MD 21218, USA.

Proceedings of the National Academy of Sciences of the United States of America
|March 16, 2007
PubMed
Summary

Repeat proteins unfold cooperatively due to uniform stability. Destabilizing mutations reveal a threshold where repeats decouple, impacting protein folding dynamics.

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Area of Science:

  • Protein folding and biophysics
  • Structural biology
  • Molecular dynamics

Background:

  • Repeat proteins feature tandem structural modules with local contacts.
  • Despite local contacts, these proteins often unfold as a single, coupled unit.
  • Previous studies suggest uniform stability distribution contributes to cooperative unfolding of Notch ankyrin domain.

Purpose of the Study:

  • To investigate the role of energy landscape uniformity in cooperative unfolding.
  • To quantify the destabilization threshold for repeat decoupling in Notch ankyrin domain variants.
  • To understand how uneven energy landscapes affect protein unfolding intermediates.

Main Methods:

  • Generated 15 Notch ankyrin domain variants with destabilizing substitutions.

Related Experiment Videos

  • Applied free energy additivity analysis to quantify coupling limits.
  • Utilized m-value analysis and differential unfolding transitions (CD vs. fluorescence).

    • Main Results:

    • Quantified a free energy coupling limit of approximately 4 kcal/mol for repeat decoupling.
    • Observed that increasing energy landscape unevenness leads to repeat decoupling.
    • Experimental data quantitatively matched predictions based on energy landscape analysis.

    Conclusions:

    • A uniform distribution of local stability is crucial for cooperative unfolding of repeat proteins.
    • Destabilizing mutations can lead to decoupling of structural modules.
    • The study provides quantitative insights into the relationship between energy landscape and protein folding cooperativity.