Chemical synthesis of proteins in solution

Development of a novel strategy suitable for the solution synthesis of proteins is described, wherein the entire molecule is assembled from fully protected segments in the size range of about 10 residues. Each segment is designed so as to have a common structure of Boc-peptide-OPac (Pac: phenacyl) and all of the side-chain functional groups are protected by Bzl-based groups. New types of solvent systems are described for dissolving fully protected segments in which the segment condensation reactions in solution can be carried out smoothly. After removal of the Boc or Pac group, the segments are coupled together to obtain the entire sequence using the 1-ethyl-3-(3'-dimethylaminopropyl)-carbodiimide/3, 4-dihydro-3-hydroxy-4-oxo-1,2,3-benzotriazine method. The side-chain protecting groups are then removed by HF and the liberated peptide is subjected to folding reactions to obtain the native conformation. Applying the strategy, the 123-residue human angiogenin, the 121-residue human midkine, the 136-residue human pleiotrophin, and the 238-residue Aequoria green fluorescent protein were synthesized successfully.