The Proteomics Core Facility at Rikshospitalet HF
The Proteomics Core Facility at Rikshospitalet HF
The Proteomics Core Facility (PCF) at Oslo University Hospital – Rikshospitalet provides proteomic service to all research groups in the HSØ region in need of it. We are constantly expanding our service portfolio to serve the increasing demands for such analysis. New techniques and expertise are made available as service regularly.
In 2012, the PCF had further extended its proteomic service provided to biomedical research groups in the HSØ region. PCF eye witnessed an increase in service demand, and this together with its outdated instrumentation resulted in PCF high priority to acquire instrumentation upgrade. In April 2012 PCF acquired a quadrupole-Orbtrap QExactive instrument (Thermo) to increase its competitiveness. This instrument is nowadays considered one of the top in the market. With the arrival of this new high-end instrument, users demand and interest increased accordingly as well. The number of runs performed from April to December surpassed 2000 runs, more than what was performed from 2008 until 2011.
In addition, due to the end of the FUGE program in 2011, most of the current available funding for PCF is almost exclusively from HSØ. However, in early 2012 PCF received a 1-year grant from a bilateral FUGE program for a project regarding establishment of high-resolution mass spectrometry in Norway. In this project PCF established collaboration with the Max Planck Institute for Biochemistry in Martienried, directed by Matthias Mann, a leading researcher in proteomics.
Methods that were proposed in the project and were previously implemented (see report 2011) such as the characterization of phosphorylation in peptides and the characterization of protein components in formalin-fixed paraffin-embedded biopsy clinical samples are now used at routine level. For example, PCF established collaboration with Radiumhospitalet to identify and quantify proteins from biopsied material of prostate cancer patients in order to identify cancer biomarkers. With the acquisition of high-resolution instrumentation, PCF is not only offering bioinformatics support for analysis of high-resolution mass spectrometry data together with label-free protein quantitation, but is also offering the mass spectrometry analysis itself. In the PCF current routine, it is possible to identify and quantify over 7000 protein in cell extracts from primary or immortalized cell cultures, or over 5000 proteins from paraffin-embedded tissues. This is high-quality data only available at a handful of proteomic laboratories worldwide.
Finally, in 2012 the bioinformatics position funded by NFR-FUGE and that was shared between PCF and the proteomic laboratories at the Biotechnology Centre in Oslo (Bernd Thiede) and UMB at Ås (Vincent Eijsink) was terminated. Magnus Arntzen was hired for the period, and was an active support for all groups. Together, we had published several articles in this period; therefore the bioinformatician had kept his position under different grants and is still an active support member of PCF.
Proteome Core Facility at OUH-Rikshospitalet
The Proteomics Core Facility (PCF) at Oslo University Hospital – Rikshospitalet provides proteomic service to all research groups in the HSØ region in need of it. We are constantly expanding our service portfolio to serve the increasing demands for such analysis. New techniques and expertise are now available as service. A new group leader, Gustavo de Souza, was recently hired to lead the PCF.
Recently, former group leader of PCF Burkhard Fleckenstein left the University of Oslo to follow his career in Germany. For his position, the Institute of Immunology hired Gustavo de Souza, a 33 years-old researcher with 9 years of experience in mass spectrometry and 8 years of experience in proteomics. Gustavo de Souza has a PhD in Cell Biology from the University of Sao Paulo in 2004, and after that he had two and half years of post-doctoral work at the Max-Planck Institute fur Biochemie, Martinsried, Germany. At that Institute he worked with Matthias Mann, one of the leading scientists in the field of Proteomics. In addition to expertise with the instruments currently available at the PCF, Gustavo de Souza has also knowledgeable experience with advanced instrumentation such as the LTQ-Orbitrap, and bioinformatic skills using MaxQuant for SILAC labeling and label-free quantitation analysis. Scientifically, Gustavo de Souza has a competent CV with 24 publications (23 using mass spectrometry), 14 of those in the year of 2009-2010 alone.
In addition, in 2010, the PCF had further extended its proteomic service provided to biomedical research groups in the HSØ region. The number of data analysis had surpassed 400 samples from over 36 different groups, which is as much as the total number of analysis performed in 2008 and 2009 all together. This clearly demonstrates that the PCF is a well accepted institution within the research community. And also indicates the increase in service demand that the PCF is now facing, specially regarding analysis of complex protein mixtures using Electrospray Ionization (ESI) instruments. However, it is worthy mentioning that two of three of PCF instruments are about to complete 5 years of usage, which is considered by most of the mass spectrometers manufacturers the optimal life-time of such instruments. Therefore, it is safe to assume that the PCF not only faces an increase in service demand while also faces an expected reduction in instrument performance in 2011-2012.
The ion trap mass spectrometer installed in Sept’09 which was cofunded by HSØ had severe downtime in 2010 due to troubleshooting with its liquid chromatography system. However the instrument has been stable in the last semester of the year, and since then it has been used on routine analysis more often. We are currently optimizing it to characterize phosphorylation and glycosylation in peptides. We will be soon offering such type of analysis to the community, and currently we already have the first samples being processed for characterization of glycopeptides from a group within Rikshospitalet. This is in accordance with the aims proposed in this application.
New methods have been developed and published, focusing mainly in peptide/protein pull-downs. Those helped the researchers involved in PCF to elucidate that transglutaminase 2 affinity to gluten favors celiac disease epitopes (Dørum et al., 2010), as well as binding signatures for transglutaminase 3 and 6 (Stamnaes et al., 2010). Mass spectrometry was also used to support the design of high-affinity peptides for antigen DQ2 (Jüse et al., 2010). Finally, a new method for enrichment of citrulline-containing peptides, which are important markers in rheumatoid arthritis, was also developed (Tutturen et al., 2010).
The PCF is also a node in the national-wide proteomics consortium NorProteomics (FUGE/NFR). Together with the proteomic laboratories at the Biotechnology Centre in Oslo (Bernd Thiede) and UMB at Ås (Vincent Eijsink), the PCF has received funding from FUGE-Øst for an expert in bioinformatics. This position is shared between laboratories, and Magnus Arntzen was hired for it. He was formerly an engineer at the Proteomic Unit of Bergen, and had bioinformatic collaboration with PCF current leader Gustavo de Souza (this collaboration resulted in two publications in journals ‘Bioinformatics’ and ‘Molecular and Cellular Proteomics’ in 2010). Since his arrival Magnus Arntzen has been an active member in all groups and results are already being published (Dørum et al., 2010; Arntzen et al., 2010).
Small-scale purification and mass spectrometry analysis reveal a third aquaporin-4 protein isoform of 36 kDa in rat brain.
J Neurosci Methods 2012 Oct;211(1):31-9. Epub 2012 aug 7
Evidence that HLA-DQ9 confers risk to celiac disease by presence of DQ9-restricted gluten-specific T cells.
Hum Immunol 2012 Apr;73(4):376-81. Epub 2012 jan 31
T-cell response to gluten in patients with HLA-DQ2.2 reveals requirement of peptide-MHC stability in celiac disease.
Gastroenterology 2012 Mar;142(3):552-61. Epub 2011 nov 19
Anti-PAD4 autoantibodies in rheumatoid arthritis: levels in serum over time and impact on PAD4 activity as measured with a small synthetic substrate.
Rheumatol Int 2012 May;32(5):1271-6. Epub 2011 jan 26
ApoptoProteomics, an integrated database for analysis of proteomics data obtained from apoptotic cells.
Mol Cell Proteomics 2012 Feb;11(2):M111.010447. Epub 2011 nov 8
Proteomic analysis of a multi-resistant clinical Escherichia coli isolate of unknown genomic background.
J Proteomics 2012 Mar;75(6):1830-7. Epub 2011 des 30
The preferred substrates for transglutaminase 2 in a complex wheat gluten digest are Peptide fragments harboring celiac disease T-cell epitopes.
PLoS One 2010;5(11):e14056. Epub 2010 nov 19
Gluten T cell epitope targeting by TG3 and TG6; implications for dermatitis herpetiformis and gluten ataxia.
Amino Acids 2010 Nov;39(5):1183-91. Epub 2010 mar 19
Redox regulation of transglutaminase 2 activity.
J Biol Chem 2010 Aug;285(33):25402-9. Epub 2010 jun 14
A technique for the specific enrichment of citrulline-containing peptides.
Anal Biochem 2010 Aug;403(1-2):43-51. Epub 2010 apr 23
Design of new high-affinity peptide ligands for human leukocyte antigen-DQ2 using a positional scanning peptide library.
Hum Immunol 2010 May;71(5):475-81. Epub 2010 feb 6
Substrate specificity of transglutaminases for gluten peptides
- oktober 2010
- Burkhard Fleckenstein
Exploring peptide binding to the disease associated HLA-DQ2.5 molecule by the use of peptide libraries
- september 2010
- Ludvig Sollid