Alcalase-catalysed synthesis of the precursor tetrapeptide N-benzoylarginylglycylaspartylserinamide (Bz-RGDS-NH_2) of the cell-adhesion peptide arginylglycylaspartylserine (RGDS)

摘要

In the present study, a precursor tetrapeptide Bz-RGDS-NH_2 (N-benzoylarginylglycylaspartylserinamide) of cell-adhesion peptide RGDS (arginylglycylaspartylserine) was synthesized by a novel route. First of all, the precursor tripeptide GDS-NH_2 (glycylaspartylserinamide) was synthesized by a chemical method only using aspartic acid and serine at gram scale in four steps. The linkage of the fourth amino acid Bz- Arg-OEt (N-benzoyl-L-arginine ethyl ester) to GDS-NH_2 was completed by an enzymatic method under kinetic control in water-miscible organic media. An industrial alkaline protease, Alcalase, with a wide substrate specificity, was used as the catalyst. The effects of organic solvents, pH value, reaction temperature, water content and molar ratio of substrates on the yield of Bz-RGDS-NH, synthesis were examined. The optimum reaction conditions were found to be pH 10.0, 35 ℃, 8 h, in an acetonitrile/( Na_2CO_3/NaHCO_3) buffer system (93:7, v/v) with a maximal yield of 65.2%. We found that secondary hydrolysis of the peptide product did not take place in these water-miscible organic solvents.