Abstract
Previous reports detailing mutational effects within the hydrophobic core of human acidic fibroblast growth factor (FGF-1) have shown that a symmetric primary structure constraint is compatible with a stably folded protein. In the present report, we investigate symmetrically related pairs of buried hydrophobic residues in FGF-1 (termed "mini-cores") that are not part of the central core. The effect upon the stability and function of FGF-1 mutations designed to increase primary structure symmetry within these "mini-core" regions was evaluated. At symmetry-related positions 22, 64, and 108, the wild-type protein contains either Tyr or Phe side chains. The results show that either residue can be readily accommodated at these positions. At symmetry-related positions 42, 83, and 130, the wild-type protein contains either Cys or Ile side chains. While positions 42 and 130 can readily accommodate either Cys or Ile side chains, position 83 is substantially destabilized by substitution by Ile. Tertiary structure asymmetry in the vicinity of position 83 appears responsible for the inability to accommodate an Ile side chain at this position, and is known to contribute to functional half-life. A mutant form of FGF-1 with enforced primary structure symmetry at positions 22, 64, and 108 (all Tyr) and 42, 83, and 130 (all Cys) is shown to be more stable than the reference FGF-1 protein. The results support the hypothesis that a symmetric primary structure within a symmetric protein superfold represents a solution to achieving a foldable, stable polypeptide, and highlight the role that function may play in the evolution of asymmetry within symmetric superfolds.