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P1 labels the active site Lys-210 ε-amino group of RA chemoselectively. P1 (50 μM) completely labeled RA (5 μM) within 5 min at 25 °C, as shown by LC-ESI-MS. The conjugate mass was observed at 29 847 Da (apo-RA, 29 623 Da; P1, 225 Da) (top panels). Mutation of the active site Lys-210 residue to alanine eliminated the covalent labeling of RA by P1. The unmodified RA K210A mutant mass was observed at 29 564 Da (bottom panels).
