Intramolecular Enzymatic Peptide Synthesis: Trypsin-mediated Coupling of the Peptide Bond between B22-Arginine and B23-Glycine in A Split Crosslinked Insulin

Abstract
The resealing of a split peptide bond (B22/23) in a crosslinked bovine insulin derivative by a trypsin-mediated coupling reaction in Tris buffer/glycerol/dimethylsulphoxide 1:1:1 at pH 6.5 is described. The coupling yield obtained with N.alpha.A1,N.epsilon.B29-(carbonyl-bis-methionyl) insulin [OC(Met)2] was 55-60% and reproducible yields of OC(Met)2-insulin were 40%. After removing the crosslink and purification, crystalline resynthesized insulin of high biological activity was obtained in a yield of 43% and an overall yield, based on split OC(Met)2-insulin, of 18%.

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