Propensity for C-terminal domain swapping correlates with increased regional flexibility in the C-terminus of RNase A

Authors

  • Katherine H. Miller,

    1. Biophysics Graduate Group, University of California, Berkeley, California 94720
    2. Institute for Quantitative Biosciences-Berkeley, University of California, Berkeley, California 94720-3220
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  • Susan Marqusee

    Corresponding author
    1. Biophysics Graduate Group, University of California, Berkeley, California 94720
    2. Institute for Quantitative Biosciences-Berkeley, University of California, Berkeley, California 94720-3220
    3. Department of Molecular and Cell Biology, University of California, Berkeley, California 94720
    • Biophysics Graduate Group, University of California, Berkeley, CA 94720
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Abstract

Domain swapping is a type of oligomerization in which monomeric proteins exchange a structural element, resulting in oligomers whose subunits recapitulate the native, monomeric fold. It has been implicated as a potential mechanism for protein aggregation, which provides a strong impetus to understand the structural determinants and folding mechanisms that trigger domain swapping. Bovine pancreatic ribonuclease A (RNase A) is a well-studied protein known to domain swap under extreme conditions, such as lyophilization from acetic acid. The major domain-swapped dimer form of RNase A exchanges a β-strand at its C-terminus to form a C-terminal domain-swapped dimer. To study the mechanism by which C-terminal swapping occurs, we used a variant of RNase A containing a P114G mutation that readily domain swaps under physiological conditions. Using NMR and hydrogen–deuterium exchange, we find that the P114G variant has decreased protection from hydrogen exchange compared to the wild-type protein near the C-terminal hinge region. Our results suggest that domain swapping occurs via a local high-energy fluctuation at the C-terminus.

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