Methods, setup and safe handling for anhydrous hydrogen fluoride cleavage in Boc solid-phase peptide synthesis

Muttenthaler, Markus, Albericio, Fernando and Dawson, Philip E. (2015) Methods, setup and safe handling for anhydrous hydrogen fluoride cleavage in Boc solid-phase peptide synthesis. Nature Protocols, 10 7: 1067-1083. doi:10.1038/nprot.2015.061


Author Muttenthaler, Markus
Albericio, Fernando
Dawson, Philip E.
Title Methods, setup and safe handling for anhydrous hydrogen fluoride cleavage in Boc solid-phase peptide synthesis
Journal name Nature Protocols   Check publisher's open access policy
ISSN 1750-2799
1754-2189
Publication date 2015-07-01
Year available 2015
Sub-type Article (original research)
DOI 10.1038/nprot.2015.061
Open Access Status Not yet assessed
Volume 10
Issue 7
Start page 1067
End page 1083
Total pages 17
Place of publication London, United Kingdom
Publisher Nature Publishing Group
Language eng
Subject 1300 Biochemistry, Genetics and Molecular Biology
Abstract Solid-phase peptide synthesis (SPPS) using tert-butyloxycarbonyl (Boc)/benzyl (Bzl) chemistry is an indispensable technique in many laboratories around the globe, and it provides peptides to the pharmaceutical industry and to thousands of scientists working in basic research. The Boc/Bzl strategy has several advantages, including reliability in the synthesis of long and difficult polypeptides, alternative orthogonality regarding protecting groups and ease of producing C-terminal thioesters for native chemical ligation applications. In this process, anhydrous hydrogen fluoride (HF) is used to remove the side chain protecting groups of the assembled peptide and to release the peptide from the resin, a process typically described as 'HF cleavage'. This protocol describes the general methodology, apparatus setup and safe handling of HF, with the aim of providing comprehensive information on the safe use of this valuable, well-studied and validated cleavage technique. We explain the cleavage mechanism, the physicochemical properties and risks of HF, first aid measures and the correct use of the apparatus. In addition, we provide advice on scavenger selection, as well as a troubleshooting section and video material illustrating key steps of the procedure. The protocol comprises precleavage sample preparation (30 min-2.5 h), complete HF cleavage procedure (2 h) and reaction workup (30 min).
Formatted abstract
Solid-phase peptide synthesis (SPPS) using tert-butyloxycarbonyl (Boc)/benzyl (Bzl) chemistry is an indispensable technique in many laboratories around the globe, and it provides peptides to the pharmaceutical industry and to thousands of scientists working in basic research. The Boc/Bzl strategy has several advantages, including reliability in the synthesis of long and difficult polypeptides, alternative orthogonality regarding protecting groups and ease of producing C-terminal thioesters for native chemical ligation applications. In this process, anhydrous hydrogen fluoride (HF) is used to remove the side chain protecting groups of the assembled peptide and to release the peptide from the resin, a process typically described as 'HF cleavage'. This protocol describes the general methodology, apparatus setup and safe handling of HF, with the aim of providing comprehensive information on the safe use of this valuable, well-studied and validated cleavage technique. We explain the cleavage mechanism, the physicochemical properties and risks of HF, first aid measures and the correct use of the apparatus. In addition, we provide advice on scavenger selection, as well as a troubleshooting section and video material illustrating key steps of the procedure. The protocol comprises precleavage sample preparation (30 min–2.5 h), complete HF cleavage procedure (2 h) and reaction workup (30 min).
Keyword Difficult sequences
Alpha-conotoxin
Chemical-synthesis
Deprotection
Q-Index Code C1
Q-Index Status Confirmed Code
Grant ID CTQ2012-30930
2014 SGR 137
DE150100784
254897
R01 GM098871
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2016 Collection
Institute for Molecular Bioscience - Publications
 
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