Objectives: Formation of Advanced Glycated End Products (AGEs) is a result of glycation, a non-enzymatic reaction between reducing sugars, or other α-dicarbonyl compounds, and amino groups of proteins, lipids and nucleic acids. AGEs accumulate during aging and have been implicated in the pathophysiology of numerous age-related diseases. Glycation of nuclear proteins at lysins or arginins may alter their function and may have an impact on epigenetic regulation of gene transcription.
Methods: Nuclei from human embryonic kidney cells 293A (HEK293A) and human kidney carcinoma cell line Caki-2 were isolated and AGE-modified nuclear proteins were compared via western blot using antibody against carboxymethyl-lysine (CML), hydroimidazolone MG-H1 and argpyrimidine. Nuclear Proteins were analyzed by mass spectrometry (LC-MS/MS).
Results: Different pattern of modified nuclear proteins in HEK293A and Caki-2 cell lines was observed. LC-MS/MS analysis revealed that in HEK293A cells 7 out of 3424 identified proteins are modified, while in Caki-2 cells among 2701 proteins 11 are modified. Two of the proteins, Histone H4 and Myb-binding protein 1A, were identified in both cell lines. Amount of modified peptides (12 out of 32127 identified peptides in HEK293A cells and 14 out of 23528 in Caki-2 cells) is too low for reproducible quantification of differences in these modifications. Further enrichment steps are therefore needed for reproducible results.
Conclusion: Identified AGE-modification of lysins/arginins within nuclear proteins indicate on a possibility to modify the epigenetic regulation of cells.
