engleski

High throughput screening for dipeptidyl peptidase III interacting proteins

Dipeptidyl peptidase III (DPP III), the ubiquitously expressed member of the M49 family of metallopeptidases, cleaves dipeptides from the N-termini of 3 to 10 residues long peptides. Over the last decades, DPP III was primarily investigated at the biochemical and enzymatic levels, while its physiological roles remained largely unknown. Recent advances in DPP III research showed that direct binding of DPP III to Keap1 induces translocation of the transcription factor NRF2 from cytosol into the nucleus, thus initiating the transcription of oxidative stress protective genes. The possibility that DPP III interacts with other proteins in a similar fashion prompted us to investigate the DPP III interactome by using two high-throughput screening methods. Yeast two-hybrid screening of the universal standardized human cDNA library was set up to identify direct interactors of DPP III, whereas the SILAC-based immunoprecipitation of DPP III coupled to MS analysis was envisioned to identify the DPP III-containing protein complexes in the cell. We aim to discover the novel DPP III protein interactors and to confirm these interactions by several complementary approaches, such as pull-down assays and confocal microscopy techniques. Once confirmed, novel interactors might open new directions in the investigation of the DPP III physiological roles in the future.

DPP III ; Keap1 ; oxidative stress

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