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Keywords:

  • φ-value analysis;
  • β-trefoil;
  • protein folding;
  • stability–function tradeoff;
  • protein evolution;
  • protein symmetry;
  • protein design

Abstract

The acquisition of function is often associated with destabilizing mutations, giving rise to the stability–function tradeoff hypothesis. To test whether function is also accommodated at the expense of foldability, fibroblast growth factor-1 (FGF-1) was subjected to a comprehensive φ-value analysis at each of the 11 turn regions. FGF-1, a β-trefoil fold, represents an excellent model system with which to evaluate the influence of function on foldability: because of its threefold symmetric structure, analysis of FGF-1 allows for direct comparisons between symmetry-related regions of the protein that are associated with function to those that are not; thus, a structural basis for regions of foldability can potentially be identified. The resulting φ-value distribution of FGF-1 is highly polarized, with the majority of positions described as either folded-like or denatured-like in the folding transition state. Regions important for folding are shown to be asymmetrically distributed within the protein architecture; furthermore, regions associated with function (i.e., heparin-binding affinity and receptor-binding affinity) are localized to regions of the protein that fold after barrier crossing (late in the folding pathway). These results provide experimental support for the foldability–function tradeoff hypothesis in the evolution of FGF-1. Notably, the results identify the potential for folding redundancy in symmetric protein architecture with important implications for protein evolution and design.