Proteomics /Mass spectrometry

Facility Interests:

Collaborative projects
Development of proteomics technologies
Core Facility proteomics analyses (ID, phospho-proteomics)

The Mass Spectrometry / Proteomics Facility provides protein characterization resources for the IFOM-IEO Campus scientist and outside. These services include protein identification and protein quantitation from a wide variety of sample types from simple mixtures (gel spots and bands) to complex mixtures (protein complexes, cell lysates, and plasma).

Capabilities currently offered:

• Protein Identification by nanoLC-ESI-MSMS
  Nano-LC online coupled to nanoESI-MSMS is used as standard method for reliable and high sensitive protein identification and characterization of single proteins or protein mixtures from SDS-PAGE or solution.
• Quantitative Proteomics
  Implementation of the following quantitative proteomics techniques in tandem with developing statistical analysis methods for the vast data quantities generated:
  Global protein quantitation: iTRAQ, SILAC
  Targeted protein quantitation: AQUA
• Post-translational modifications analysis
  Characterization of post-translational modifications
  Phosphorylation: detect / localize / quantitate phosposites on protein(s)
  Ubiquitin: detect / localize / quantitate
  Other PTMs – consult with us

For detailed info on Services, please go to the Facility‘s web page (please make words into a link to: http://www.ifom-ieo-campus.it/services/masspectrometry.php)

My research interests:

Glycoproteins as Cancer biomarkers

Elucidation of the genomes of different eukaryotes has shown that the increasing complexity of higher organisms is largely unrelated to the total number of genes. The human proteome contains up to 1 million proteins compared to the 25,000 genes in the genome. If differential posttranslational modifications were also to be considered, it would add orders of magnitudes to this number.
Glycosylation is one of the most abundant forms of posttranslational modifications of proteins. It is one of the most complex regulated processes of the cell, with more than 1,000 genes or more than 2% of the genome involving enzymes required for biosynthesis and metabolism of complex glycans. Complex carbohydrates play a wide array of different biological roles from direct effects on folding, processing, sorting, and functions of proteins, to serving as ligands in important adhesion events for cell-cell interactions. Most secreted proteins are glycosylated, including important tumor biomarkers such as prostate- specific antigen and the ovarian cancer marker CA125. Glycoproteins produced by cancer cells have altered glycan structures, although the proteins themselves are common, ubiquitous, abundant, and familiar. Thus, the ability to monitor changes in the glycoproteome is essential to determine possible biomarkers. Concurrently, often a disease such as cancer manifests itself in large alterations in the distribution of glycoforms presented on glycoproteins. For example, increased branching of N-linked glycans has been associated with invasion, angiogenesis and metastasis. Increased sialylation on the cell surface can promote cell detachment from primary tumors through charge repulsion, thereby inducing tumor proliferation.

Serum obtained from several types of cancers and serum from healthy subjects is to be used as the study material. The approach has two parallel facets:

• an examination of the entire glycoproteome via quantitative mass spectrometry
• global profiling of glycans from all serum glycoproteins

For the glycoproteome quantitation aspect, we plan to use the isobaric tagging method iTRAQ to detect quantitative alterations in the glycoprotein levels. For examination of total glycans, the serum glycoproteins are cleaved and preconcentrated before global glycoprofiling via IRMP or ECD.