All peptides used in this study were synthesized by the standard solid phase technique (ANYGEN Co., Ltd.). 5-Iodoacetamido fluorescein (5-IAF)-labeled peptides were prepared as previously described68 (link). Briefly, peptides containing a single cysteine residue were incubated with 5-IAF (Sigma-Aldrich) at a 1:2 molar ratio in 100 mM HEPES–NaOH pH 7.5 for 2 h at RT. The 5-IAF-labeled peptide was separated from the native peptide and excess 5-IAF by Shim-pack PREP-ODS column-equipped reversed-phase high-performance liquid chromatography (Shimadzu 10Avp). The extent of 5-IAF labeling was confirmed by mass spectrometry.
Park J.G., Kim S., Jang E., Choi S.H., Han H., Ju S., Kim J.W., Min D.S, & Jin M.S. (2022). The lysosomal transporter TAPL has a dual role as peptide translocator and phosphatidylserine floppase. Nature Communications, 13, 5851.
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