Enzyme-assisted semisynthesis of polypeptide active esters and their use

• Published in: Biochemical journal 1988-01, Vol.249 (1), p.83-88
• Main Authors: Rose, K, Herrero, C, Proudfoot, A E, Offord, R E, Wallace, C J
• Format: Article
• Language: English
• Subjects: Alanine - analogs & derivatives; Chlorophenols; Chromatography, High Pressure Liquid; Chromatography, Ion Exchange; cytochrome c; Cytochrome c Group - analogs & derivatives; Cytochrome c Group - chemical synthesis; Esters - chemical synthesis; Methods; Nitrophenols; Peptide Fragments - chemical synthesis; peptide synthesis; Peptides - chemical synthesis; Solvents; Trypsin
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/bj2490083

A method is described for the preparation of polypeptides activated uniquely at the C-terminus. The polypeptide is incubated in a concentrated solution of an amino acid active ester, the latter having its amino group free but adequately protected by protonation. The amino acid ester is coupled via its amino group to the C-terminus of the polypeptide by enzymic catalysis (reverse proteolysis). The resulting polypeptide C-terminal active ester is then isolated and coupled to a suitable amino component (generally a polypeptide) in a subsequent chemical coupling. The method appears to be generally applicable; fragments of horse heart cytochrome c, and porcine insulin, are used as examples. Two new analogues of cytochrome c have been prepared by using this method, with yields of up to 60% in the final coupling. Scope and limitations of the method are discussed.