The protein chemistry facility (PCF) is a research and service platform of IBAR performing protein, peptide, proteome and subproteome analysis.
The protein chemistry facility (PCF) is a research and service platform of IBAR performing protein, peptide, proteome and subproteome analysis. The facility was established in 2012 by a EU funded grant PROCERA (POSCCE-A2-O2.2.1-2009-4) in the Department of Bioinformatics and Structural Biochemistry (IBAR-DBSB).
Its main research topics are closely related with the focus-area of other groups and departments of IBAR: proteome and subproteome characterization in melanoma, protein-protein interactions in biological systems, co- and post-translational modification analysis with a special focus on glycosylation, imunopeptidome investigation, identification of specific protein interactors. Another priority is the study of protein phosphorylation, a dynamic process that implies the complementary activities of protein kinases and protein phosphatases. Mass spectrometric analysis of the samples implies the use of CID, HCD and ETD fragmentation methods. Data analysis and interpretation of results is made using Thermo Scientific™ Proteome Discoverer™ software. For phosphosite localization, besides the automated analysis (PhosphoRS Node in Proteome Discoverer) a manual inspection of the raw tandem mass spectra of the peptide is often made.
As a service unit the facility can provide the following methods of analysis:
Thermo Scientific Easy-nanoLC II:
The Easy-nanoLC is a binary pump system able to separate biomolecules on capillary columns at low flows (100-1000 nL/min). It is used to separate peptides on reversed-phase C18 nano columns.
Thermo Scientific LTQ-Orbitrap Velos Pro:
This is a hybrid mass analyzer composed of a fast linear ion trap (up to 12.5 Hz) and a high resolution Orbitrap analyzer (up to 100 000 resolution at m/z 400) with mass accuracy in the low ppm range. It is used for peptide, protein, small molecule and other biomolecules analysis.
Thermo Scientific Accela 600:
This separation instrument is equipped with a 600 bar pump allowing small molecule and biomolecule routine analysis, but also applications requiring high flow separations (up to 5 ml/min). It is connected to two detection systems: an UV-VIS PDA detector and a fluorescence detector (Thermo Finnigan Surveyor FL Plus Detector). It is used for peptide fractionation and purification.
Thermo Scientific Dionex IC5000:
Dionex IC5000 is an instrument dedicated to ion chromatography. It features a quaternary pump, an electrochemical detector and an additional UV detector (also includes a desalting device) for to the analysis of various ions. It is dedicated to the to the separation highly polar molecules, like amino acids, glycans, hydrogen peroxide or nitrates.
As a research platform the members of the facility actively participate in the elaboration and optimization of new methods and protocols for protein analysis in biological systems. Furthermore the team also contributes to the elaboration of grant applications and research publications that include the expertise of its members.
Selected research grants that included the participation of IBAR-PCF members:
Selected publications that included the participation of IBAR-PCF members: