Training Courses

Thursday, June 28th & Friday, June 29th

9.00-12.30h / 13.30-16.00

Workshop I - Utilizing Nano UHPLC to Maximize Peptide and Protein Identification Results, Steffen Liedtke & Evert-Jan Sneekes &Remco Swart

Dionex now part of Thermo Fisher Scientific

Thursday

In the theory session the following topics are covered: introduction nano LC, peak capacity, optimization of LC of peptides, coupling aspects of nano LC/MS. In addition some common diagnostics and troubleshooting will be presented.

The hands-on session covers the following topics: installation of nano LC columns, working with nanoViper, programming optimal gradient conditions, reviewing high resolution nano LC data.

 

Workshop II – High Resolution 2D LC of Proteins and Peptides, Steffen Liedtke & Evert-Jan Sneekes &Remco Swart

Dionex now part of Thermo Fisher Scientific

Friday

In the theory session the following topics are covered: introduction PS-DVB monolithic columns, challenges in protein separations, optimization of LC of proteins, fractionation capabilities, on-line and off-line 2D LC techniques.

The hands-on session covers the following topics: installation of monolithic columns, working with nanoViper, programming optimal gradient conditions, reviewing high resolution protein separations.

Thursday, June 28th & Friday, June 29th

Gel-free Proteomics: Mastering MS Data, Marc Vaudel, Lennart Martens & Albert Sickmann 

Hands-On Workshop
9.00-12.30h / 13.30-16.00

Gel-free Proteomics Data Interpretation: Beautiful Experiments deserve more than the Mascot Summary

Gel-free proteomics have the characteristic of producing vast amounts of data: current experiments typically generate hundreds of thousands of spectra themselves containing hundreds of peaks. Before obtaining results from your experiment, it is thus mandatory to conduct a thorough statistical analysis of the data. Hopefully, methods and tools were – and are still being – developed in order to make this task feasible. Moreover, we benefit from numerous online resources making available the proteomic knowledge accumulated along the years.

Yet, it is not straightforward to apply these methods in everyday workflows. Even worse, their bad application can simply lead to drawing false scientific conclusions. Also, external resources are often unused, simply because they are unknown. These issues will be addressed in the bioinformatics for proteomics session where we will go through some of the main topics in proteomics data processing in an interactive and user-friendly mode.
The workshop will cover the following themes:

  • Processing of raw files
  • MS/MS spectra identification
  • Proteins and Peptides identification and validation
  • PTM analysis with a focus on phosphorylation
  • Sequence pattern recognition using IceLogo
  • Quantification results interpretation: who is regulated?
  • Online repositories, submission to PRIDE and browsing online experiments
  • Use of external resources for biological interpretation:

a.) 3D structural analysis with a focus on phosphorylation location on the structure
b.) Accessing UniprotKB protein information
c.) Pathway analysis
d.) Interaction analysis
e.) Gene Ontology analysis of the obtained results

Organizers:
Lennart Martens, Ghent University
Marc Vaudel, ISAS Dortmund 

Thursday, June 28th & Friday, June 29th 

 Phosphorylation Analysis, Rene Zahedi

Hands-On Workshop
9.00-12.30h / 13.30-16.00 

In this hands-on workshop, 4-5 partipants will process samples for phosphoproteomic analyses.

Cell lysates will be proteolytically digested and processed, phosphopeptides will be enriched and analyzed by nLC-MS/MS with high mass accuracy. Moreover, participants will learn about current quality control issues in phosphoproteomics such as correct site localization, false discovery rates, and will be introduced to phosphopeptide spectrum interpretation. In addition, quantitative phosphoproteomics strategies, such as iTRAQ, SILAC and SRM will be covered.

Topics:

  • nLC MS/MS analysis and processing of raw files
  • phosphopeptide enrichment
  • quantitative phosphoproteomics
  • data analysis and pitfalls during spectrum interpretation
  • troubleshooting
  • Quantification results interpretation: who is regulated?

Thursday, June 28th & Friday, June 29th

Thiol Redox-Proteomics small pdf icon
Lars Leichert, Alexandra Müller

Hands-On Workshop
9.00-12.30h / 13.30-16.00

This hands-on workshop will cover the general principles of MS-based redox-proteomics using a model protein. The workshop is aimed at proteome-researchers with an interest in oxidative stress and redox-biology.

Oxidative stress is the inevitable consequence of an aerobic life-style. Reactive oxygen species, the unwanted by-products of many essential cellular processes, such as respiration, can damage virtually all important bio-molecules. Therefore it is not surprising that many pathological conditions are associated with oxidative stress and that on a molecular level reactive oxygen species are thought to be the causative agent of cellular ageing.

While these destructive properties of oxidative stress have long been known, more recently it became apparent, that so-called redox-regulated proteins are not damaged by oxidative stress, but instead use the amino-acid cysteine as a nano-switch to sense the redox-environment of the cell.

In the last few years quantitative proteomic methods have been developed to identify these redox-sensitive cysteines and to study cysteine oxidation under pathological and physiological stress conditions on a global level. Here, we will cover the basic principles of these methods and test our newly gained knowledge on a redox-regulated model protein.

The workshop will cover the following topics:

  • Oxidative thiol modifications, theory and practice
  • The pitfalls of redox-proteomics and how to avoid them best
  • Differential trapping of reduced and oxidized thiols
  • Isotope labeling of proteins for MS based quantification of the redox state
  • Correlation of the MS data with biological activity

This course is limited to 6 participants.

Organizers:
Lars Leichert, Medizinisches Proteom-Center, Bochum
Alexandra Müller, Medizinisches Proteom-Center, Bochum