Projekter pr. år
Protein phosphorylation and cell signaling networks have been studied thoroughly by large-scale phosphoproteomics technologies, including affinity enrichment methods and LC-MS/MS. A vast majority of studies reported phosphorylation of Ser, Thr and Tyr residues in proteins based on immobilized metal affinity chromatography, i.e. IMAC and TiO2 (pS, pT, pY) and antibody-based enrichment (pY). Recently, we reported the application of molecularly imprinted polymers (MIPs) for pY-peptide enrichment (Bllaci et al., 2017). Here, we report initial results obtained by using a novel monolithic MIP capillary column directed towards phosphohistidine (pHis) residues and a second MIP for targeting specific pY motifs. Phosphohistidine residues are labile at acidic conditions necessitating special precautions during sample preparation and LC-MS analysis. We used chemical phosphorylation using potassium phosphoramidate (Wei et al. 1991) to generate myoglobin pHis-peptides, which were used for testing the crushed monolith and column MIP by LC-MS/MS. Peptides from ZAP70 kinase were spiked on a 12 proteins digest for characterizing the pY-motif specific MIP. This method was performed on batch mode with different spiking levels to address the sensitivity of the MIP by LC-MS/MS analysis of the elution fractions. We will present this new modification-specific proteomics strategy and preliminary results obtained by using MIPs in combination with LC-MS/MS for biomolecule characterization.
|Status||Udgivet - 2019|
|Begivenhed||Proteomics in Cell Biology and Disease Mechanisms: EMBL–Wellcome Genome Campus Conference - EMBL Advanced Training Centre, Heidelberg, Tyskland|
Varighed: 7. mar. 2019 → 9. mar. 2019
|Konference||Proteomics in Cell Biology and Disease Mechanisms|
|Lokation||EMBL Advanced Training Centre|
|Periode||07/03/2019 → 09/03/2019|
EU Horizon2020 - Excellence Science - Marie Curie - Innovative Training Networks - ETN: Smart capture phases for proteomics, glycomics and biomarker assays
01/11/2016 → 31/10/2020
Projekter: Projekt › Forskning
Arribas Díez, I., Anil Incel, & Chau Huynh (2019). Molecularly Imprinted Polymers for characterization of rare phosphorylation events. Poster session præsenteret på Proteomics in Cell Biology and Disease Mechanisms, Heidelberg, Tyskland.