Enzymatic synthesis of polypeptides in natural deep eutectic solvents

We are first to report the protease-catalyzed polypeptide synthesis in the natural deep eutectic solvent (DES) choline chloride/glycerol (ChCl:Gly) at a 1:2 molar ratio with 30 vol% of water using subtilisin Carlsberg (alkaline protease from Bacillus licheniformis). Poly-L-phenylalanine, poly-a-L-lysine, and the co-polypeptides poly(L-phenylalanine-a-L-lysine) as model polypeptides are produced from amino acid ester substrates in 40–70% yield and molecular weights ranging from 5 to 14 KDa according to dynamic light scattering (DLS). The natural DES is composed of 30 vol% water, whereas no polypeptides are formed at lower water contents. Poly-L-phenylalanine displays supramolecular self-assembly into homogenous nanotubes in water/isopropyl alcohol mixtures. This enzymatic route to polypeptides has advantages over previous reports in water and polar compressed fluids because of the higher molecular weights and greener process, respectively.