Synthesis of FRET-HA Peptide Probe

The FRET-HA peptide (7-MCA-GLFGAIAGFIE-GMVDGWYGK(DNP)rrr-NH₂) was synthesized on a solid support using the Fmoc/tBu strategy on a rink amide resin as previously reported^{39 (link)}. Briefly, the Fmoc group was removed with 20% piperidine in dimethylformamide, and chain elongation was achieved by standard HBTU/HOBt chemistry using three equivalents of protected amino acids or 7-MCA. After synthesis completion, the protected peptide resin was treated with a TFA/phenol/H₂O/thioanisole/1,2-ethanedithiol (82.5:5:5:2.5, v/v/v/v) mixture. The crude material obtained was purified using HPLC. The structures of the FRET peptides were confirmed using MS.