Learning about peptides
Research, protocols and updates on peptides for scientific use.
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Click chemistry in peptide research: azide-alkyne cycloaddition for targeted bioconjugation
What makes a reaction "click chemistry"? Click chemistry is a collective term for a class of chemical reactions that proceed rapidly, selectively and under mild conditions, with high yield and few unwanted...
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Peptide mapping of research peptides: enzymatic digestion and LC-MS/MS for sequence confirmation
An intact mass determination by LC-MS confirms that a research peptide has the correct molecular weight, but says little about the exact order of the amino acids within it. Two peptides with...
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Green solvents in SPPS: why DMF is under pressure and what the alternatives are
Dimethylformamide (DMF) has for decades been the standard solvent in Fmoc-based solid-phase peptide synthesis (SPPS): it swells polystyrene and PEG resins well, readily dissolves protected amino acids and coupling reagents, and is...
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Residual solvents in research peptides: what GC analysis reveals after lyophilisation
After solid-phase synthesis, cleavage, preparative HPLC purification and lyophilisation, a research peptide is rarely completely free of the chemicals that made the synthesis and purification process possible. Besides the water content, which...
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Stapled peptides: how a hydrocarbon “staple” improves secondary structure and protease resistance in research
Linear peptides have a structural problem: outside a folded protein, short alpha-helical sequences lose their shape as soon as they go into solution. For researchers who want to study a specific helix...
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Stable isotope labelled peptides: internal standards for quantitative LC-MS/MS
Quantitative mass spectrometry plays an ever larger role in peptide-related research, from proteomics studies to the characterisation of synthetic research peptides. But an LC-MS/MS signal is not in itself a reliable measure...
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Disulfide bridges in multi-cysteine research peptides: oxidative folding and regioselective strategies
Many bioactive research peptides derive their three-dimensional structure not only from their amino acid sequence, but also from one or more disulfide bridges between cysteine residues. In peptides with only a single...
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Native Chemical Ligation: Beyond The Length Limit Of SPPS
Why SPPS does not scale indefinitely Solid-phase peptide synthesis (SPPS) is the standard method for building research peptides, residue by residue, on a solid resin support. Earlier articles on this blog covered...
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Residual TFA after counterion exchange: what ion chromatography reveals
Why residual TFA matters Trifluoroacetic acid (TFA) is indispensable in the preparative HPLC purification of research peptides: it acts as an ion-pairing reagent and produces sharp, well-separated peaks during purification. After counterion...
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Capillary electrophoresis as an orthogonal technique alongside RP-HPLC for research peptides
When a research peptide has come through preparative HPLC purification and counterion exchange, a crucial question remains: is the reported purity percentage actually correct? RP-HPLC is the industry standard, but no separation...
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From pure fraction to end product: pooling and counterion exchange after preparative HPLC
Preparative HPLC yields a series of fractions, each with its own purity profile. But the fraction coming out of the collector is not yet an end product. Between "pure peak" and "documented...
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Preparative HPLC purification of research peptides: from crude mixture to pure fraction
After the cleavage step there is no pure research peptide on the bench, but a complex crude mixture. The previous article in this series covered how scavengers during TFA cleavage prevent reactive...
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