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Peptide thioester preparation by Fmoc solid phase peptide synthesis for use in native chemical ligation

journal contribution
posted on 2000-05-01, 00:00 authored by A B Clippingdale, Colin BarrowColin Barrow, J D Wade
Established methodology for the preparation of peptide thioesters requires the use of t-butoxycarbonyl chemistry owing to the lability of thioester linkers to the nucleophilic reagents used in Fmoc solid phase peptide synthesis. Both the greater ease of use and the broad applicability of the method has led to the development of an Fmoc-based methodology for direct peptide thioester synthesis. It was found that successful preparation of a peptide thioester could be achieved when the non-nucleophilic base, 1,8-diazabicyclo[5.4.0]undec-7-ene, together with 1-hydroxybenzotriazole in dimethylformamide, were used as the N(alpha)-Fmoc deprotection reagent. Native chemical ligation of the resulting thioester product to an N-terminal cysteine-containing peptide was successfully performed in aqueous solution to produce a fragment peptide of human alpha-synuclein. The formation of aspartimide (cyclic imide) in a base-sensitive hexapeptide fragment of scorpion toxin II was found to be significant under the deprotection conditions used. However, this could be controlled by the judicious protection of sensitive residues using the 2-hydroxy-4-methoxybenzyl group.

History

Journal

Journal of peptide science

Volume

6

Issue

5

Pagination

225 - 234

Publisher

Wiley

Location

Chichester, Eng.

ISSN

1075-2617

Language

eng

Publication classification

C1.1 Refereed article in a scholarly journal

Copyright notice

2000, European Peptide Society and John Wiley & Sons, Ltd.