Convergent solid-phase synthesis of hirudin

✍ Scribed by Spyros Goulas; Dimitrios Gatos; Kleomenis Barlos

• Book ID: 105360509
• Publisher: John Wiley and Sons
• Year: 2006
• Tongue: English
• Weight: 170 KB
• Volume: 12
• Category: Article
• ISSN: 1075-2617
• DOI: 10.1002/psc.691

✦ Synopsis

Abstract

Hirudin variant 1 (HV1), a small protein consisting of 65 amino acids and three disulfide bonds, was synthesized by using Fmoc‐based convergent methods on 2‐chlorotrityl resin (CLTR). The linear sequence was assembled by the sequential condensation of 7 protected fragments, on the resin‐bound 55–65 fragment. The conditions of fragment assembly were carefully studied to determine the most efficient synthetic protocol. Crude reduced [Cys^16, 28^(Acm)]‐HV1 thus obtained was easily purified to homogeneity by RP‐HPLC. Disulfide bridges were successfully formed by a two‐step procedure, involving an oxidative folding step to form Cys^6^–Cys^14^ and Cys^22^–Cys^39^ linkages, followed by iodine oxidation to form the Cys^16^–Cys^28^ bond. The correct disulfide bond alignment was established by peptide mapping using Staphylococcus aureus V8 protease at pH 4.5. Copyright © 2005 European Peptide Society and John Wiley & Sons, Ltd.