The discovery of labile methyl esters on proliferating cell nuclear antigen by MS/MS

The post‐translational modification of proliferating cell nuclear antigen (PCNA) has been implicated in modulating its function for over 20 years. With multiple interacting partners, PCNA is involved in processes ranging from DNA replication and repair to cell cycle control and apoptosis. The ability of PCNA to distinguish between specific binding partners in different tasks is currently of intense interest, and several post‐translational modifications have been reported to modulate its function. Unfortunately, these reports have produced contradictory information on the type(s) of modification present on the molecule. Here we report a detailed structural analysis of a single acidic PCNA isoform, cancer‐specific polyferating nuclear anitgen (csPCNA), isolated from breast cancer cells by 2D‐PAGE and LC‐MS/MS. With this approach we fully characterized the csPCNA isoform and confidently identified a single post‐translational modification, methyl esterification. Interestingly, the methyl esters consistently localized to 15 specific glutamic and aspartic acid residues of csPCNA. The methyl esterification of csPCNA represents a novel type of post‐translational modification in mammalian cells that could ultimately hold the key towards unlocking its diverse functions.