Update information. Mutations in the human protein DJ-1/PARK7 are found to be involved in familial Parkinson’s disease. However, the molecular mechanism of the protein remains unclear. Lately, DJ-1 was found to prevent the acylation of amino groups in proteins. The process of acylation requires a cyclic 3-phosphoglyceric anhydride (cPGA). The cPGA has been shown to be an intermediate product of 1,3-biphosphoglycerate and to be destroyed by DJ-1’s hydrolase activity. In our study, we synthesized and characterized cPGA. Furthermore, we demonstrated an endogenous catalytic hydrolase activity of DJ-1 towards cPGA in crude protein extracts of human HCT116 cells. The addition of synthesized cPGA to DJ-1 knockout HCT116 cytoplasmic cell lysate caused a dramatic increase in phosphoglycerate-modified proteins. The submitted results contain proteomic analysis of phosphoglycerate modifications on lysine residues in wild type and DJ-1 knockout cytoplasmic proteins of HCT116 cells when cPGA was added. Our study confirms that DJ-1 prevents the formation of phosphoglycerate modifications on amino groups of proteins.