Externt MBB seminarium: Dr. Karl Mechtler "The Dark Matter of the Proteome"
Information följer på engelska, eftersom seminariet hålls på engelska.
Seminar with Dr. Karl Mechtler, Head of Protein Chemistry Facility at Research Institute of Molecular Pathology (IMP), Vienna, Austria.
"The Dark Matter of the Proteome"
In shotgun proteomics, proteins are enzymatically digested into peptides that are fed into a highly automated pipeline for chromatography and mass spectrometry (MS) analysis.
When applied to complex mixtures of proteins from cells or tissues, the technique yields tens thousands of peptide sequences, only a minority of which can be mapped to their proteins of origin.
Despite the efforts of many researchers, the identity and origin of the unassigned peptides have remained a mystery. Most dark proteins are short, rarely interact with other proteins, and are:
- Frequently excreted and only have a small number of evolutionary relatives
- Sample preparation (e.g. non-soluble membrane proteins
- Unexpected post-translational modifications
- Protein of interest is not in database (long noncoding RNAs, SNPs, ...)
Fragmentation behavior of peptides and their modifications – more than 50 % of “dark” MS/MS spectra
On of this unexpected post-translational modification is glycosylation. More than half of all human proteins are glycosylated. This covalent attachment of carbohydrates can alter protein activity. However, existing methods for studying the glycoproteome typically requires laborious sample processing and proprietary software. They also often require enzymatic removal of the glycan, which makes it difficult to identify attachment sites. Recently we developed a proteomic method to identify intact glycopeptides in proteomics data. Focusing on the intact fragments allows both the structure of the glycan and the attachment site in the associated protein to be identified.