Custom Peptide Synthesis: From Sequence to Shipment in 5–10 Business Days
Peptide Synthesis Technology Overview
Modern solid-phase peptide synthesis (SPPS), pioneered by R. Bruce Merrifield (Nobel Prize, 1984), remains the gold standard for custom peptide production. Boston Molecules employs Fmoc/tBu chemistry on automated synthesizers (CEM Liberty Blue and Biotage Initiator+), enabling routine synthesis of peptides up to 100+ residues with consistent quality.
Synthesis Methodology
Standard Protocol
Each synthesis cycle consists of:
- Fmoc deprotection — 20% piperidine in DMF, 2 × 5 min at RT or 1 × 1 min at 90°C (microwave)
- Amino acid coupling — 4 eq Fmoc-AA-OH, 4 eq HATU, 8 eq DIPEA in DMF, 5–10 min at 75°C
- Capping (optional) — Ac₂O/DIPEA to block unreacted sites and prevent deletion sequences
- Washing — 3 × DMF between steps
For difficult sequences (polyproline, β-sheet aggregators), we employ pseudoproline dipeptides, backbone protection (Dmb/Hmb), and chaotropic additives (0.8M LiCl) to disrupt on-resin aggregation.
Modification Capabilities
| Modification | Chemistry | Application |
|---|---|---|
| Phosphorylation (pSer, pThr, pTyr) | Pre-phosphorylated building blocks | Kinase substrate studies, signaling |
| Biotinylation | N-terminal Biotin-NHS or Biotin-PEG₄ | Pull-down assays, ELISA |
| FITC/FAM labeling | N-terminal FITC-Ahx or 5-FAM | Cell permeability, imaging |
| PEGylation | Mini-PEG₂₋₁₂ spacers | Solubility, pharmacokinetics |
| Isotope labeling | ¹³C, ¹⁵N Fmoc-AAs (Cambridge Isotope) | NMR, quantitative mass spec |
| D-amino acid substitution | Fmoc-D-AA-OH building blocks | Proteolytic stability, retro-inverso |
| Acetylation / Amidation | Ac₂O (N-term), Rink amide resin (C-term) | Charge neutralization, stability |
Quality Control Pipeline
Every peptide undergoes a rigorous three-step QC process:
- Analytical RP-HPLC — C18 column, 0.1% TFA/ACN gradient, UV detection at 214 nm and 280 nm. Purity reported as % of main peak area.
- Mass Spectrometry — MALDI-TOF (Bruker Autoflex) for peptides <5 kDa; ESI-MS (Thermo Q Exactive) for larger peptides and accurate mass determination (±0.01 Da)
- Amino Acid Analysis (optional) — acid hydrolysis followed by derivatization and HPLC quantitation for precise concentration determination
Turnaround and Pricing
| Peptide Type | Purity | Turnaround | Scale |
|---|---|---|---|
| Standard linear (<30 AA) | Crude / >70% / >95% / >98% | 5–7 business days | 1–100 mg |
| Long peptide (30–80 AA) | >90% / >95% | 10–15 business days | 1–20 mg |
| Modified peptide | >95% | 7–12 business days | 1–50 mg |
| Cyclic / Stapled | >95% | 12–20 business days | 1–10 mg |
| Peptide library (96-well) | Crude | 10–15 business days | 1–5 mg each |
References
- Merrifield, R.B. “Solid Phase Peptide Synthesis. I. The Synthesis of a Tetrapeptide.” Journal of the American Chemical Society 85, 2149–2154 (1963)
- Coin, I. et al. “Solid-phase peptide synthesis: from standard procedures to the synthesis of difficult sequences.” Nature Protocols 2, 3247–3256 (2007)
Leave a Reply