Fmoc-Based Synthesis of Peptide-Thioesters: Application to the Total Chemical Synthesis of a Glycoprotein by Native Chemi
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- 作者：Youngsook Shin ; Katharine A. Winans ; Bradley J. Backes ; Stephen B. H. Kent ; Jonathan A. Ellman ; and Carolyn R. Bertozzi
- 刊名：Journal of the American Chemical Society
- 出版年：1999
- 出版时间：December 22, 1999
- 年：1999
- 卷：121
- 期：50
- 页码：11684 - 11689
- 全文大小：135K
- 年卷期：v.121,no.50(December 22, 1999)
- ISSN：1520-5126
文摘
The technique of native chemical ligation has enabled the total chemical synthesis of proteins withmolecular weights far in excess of those achievable by conventional stepwise solid-phase peptide synthesis.The method involves the condensation of two unprotected peptide segments, one bearing a C-terminal 
thioesterand the other an N-terminal cysteine residue, to afford a protein with a native amide linkage at the site ofligation. Here we report an extension of the native chemical ligation method to the total synthesis of aglycosylated protein, the antimicrobial O-linked glycoprotein diptericin. The major challenge in our synthesiswas preparation of a 24-residue glycopeptide-thioester segment, which was complicated by the incompatibilityof glycosidic linkages with Boc chemistry and by the incompatibility of thioesters with Fmoc chemistry. Theuse of an alkanesulfonamide "safety-catch" linker circumvented this problem and permitted the solid-phasesynthesis of the glycopeptide-thioester using standard Fmoc chemistry protocols. Ligation of this thioesterwith a 58-residue glycopeptide bearing an N-terminal cysteine residue yielded the full-length glycoproteinwith two sites of glycosylation. The fully deprotected diptericin glycoform was active in antimicrobial assays.
thioesterand the other an N-terminal cysteine residue, to afford a protein with a native amide linkage at the site ofligation. Here we report an extension of the native chemical ligation method to the total synthesis of aglycosylated protein, the antimicrobial O-linked glycoprotein diptericin. The major challenge in our synthesiswas preparation of a 24-residue glycopeptide-thioester segment, which was complicated by the incompatibilityof glycosidic linkages with Boc chemistry and by the incompatibility of thioesters with Fmoc chemistry. Theuse of an alkanesulfonamide "safety-catch" linker circumvented this problem and permitted the solid-phasesynthesis of the glycopeptide-thioester using standard Fmoc chemistry protocols. Ligation of this thioesterwith a 58-residue glycopeptide bearing an N-terminal cysteine residue yielded the full-length glycoproteinwith two sites of glycosylation. The fully deprotected diptericin glycoform was active in antimicrobial assays.