Zn(II)-DPA functionalized graphene oxide two-dimensional nanocomposites for N-phosphoproteins enrichment

Protein N-phosphorylation is increasingly recognized as a critical post-translational modification in central metabolism and cell signaling. However, other functions of N-phosphorylation in organism are largely unexplored which is attributed to lack high efficient enrichment methods. To address this, Bis(zinc(II)-dipicolylamine) functionalized graphene oxide nanocomposites (Zn(II)-DPA-GO) was designed via loading bis(zinc(II)-dipicolylamine) (Zn(II)-DPA) on graphene oxide surface by π-π stacking, with the advantages of large immobilization amount of Zn(II)-DPA, layered structure, excellent solubility and low non-special adsorption. The material was firstly verified excellent enrichment ability for standard proteins and E.coli lysates. By integrating the enrichment method into proteomics workflow, 36 N-phosphoproteins functioned in metabolic process, biological regulation, localization, cellular processes were identified from Bacillus subtilis. Furthermore, motif analysis showed that leucine and isoleucine were enriched near the sites, providing potential clues for the discovery of new sites. The method provides an alternative for the discovery of N-phosphoproteins with significant biological functions.