Hairpin peptides are synthesized on either an Applied Biosystem 433A or Advanced ChemTech 496 synthesizer employing standard Fmoc (9-fluorenyl methoxycarbonyl) solid-phase peptide synthesis methods and purified using RP-HPLC, using C18 and/or C8 stationary phases and a water (0.1% TFA)/acetonitrile(0.085% TFA) gradient as previously described.(18 (link),20 (link)) The resins used for the synthesis were Wang resins preloaded with the C-terminal amino acid. Peptides were cleaved from the resin using a 95:2.5:2.5 trifluoroacetic acid (TFA): triisopropylsilane: water mixture. Full-length WW domains were ordered from GenScript as the crude products for subsequent in-house HPLC purification. The sequences of all peptides were confirmed by the molecular ions observed using a Bruker Esquire ion-trap mass spectrometer.
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Synthesis and Purification of Hairpin Peptides
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Acetonitrile
Amino acid
Hplc
Peptides
Resin
Solid phase peptide synthesis methods
Synthesis
Trifluoroacetic acid
Wang resins
Ww domains
Corresponding Organization :
Other organizations : University of Washington
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Variable analysis
independent variables
- Synthesis method (Applied Biosystem 433A or Advanced ChemTech 496 synthesizer)
- Solid-phase peptide synthesis using Fmoc chemistry
- Purification method (RP-HPLC using C18 and/or C8 stationary phases)
- Gradient used for RP-HPLC (water (0.1% TFA)/acetonitrile (0.085% TFA))
- Purified hairpin peptides
- Purified full-length WW domains
- Resin used for solid-phase synthesis (Wang resins preloaded with the C-terminal amino acid)
- Cleavage conditions (95:2.5:2.5 trifluoroacetic acid (TFA): triisopropylsilane: water mixture)
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