Fig. 1.

Novel modifications of active site cysteine detected in GAPDH. A, Summary of cysteine modifications observed in cellular GAPDH purified from immuno-precipitation of HEK293 cells, by tandem MS. Peptide m/z and calculated mass, and detected nominal mass changes at 152C/156C and 247C residues were presented. B, MS/MS spectra of active site “CXXXC” containing unknown mass shift +64 and −16 Da. C^aa indicates acrylamide adduct of Cys residue and star indicates fragment ion that lost H₂O or NH₃. Presumed chemical structure of each mass shift at Cys residue was presented on the right side. C, The immuno-precipitated GAPDH was separated on 2D-PAGE and detected with Coomassie staining. D, Quantitative analyses of modifications in peptide ¹⁴⁶IISNASCTTNCLAPLAK¹⁶² based on precursor ion intensities in supplemental Fig. 1B using Glu-fibrinopeptide as an internal standard.