Frank Siedler - Mass Spectrometry

Project Group "Mass Spectrometry for Archaeal Systems Biology"

Halophilic archaea are perfect model organisms for the analysis in a systems biology context. Based on genome sequences determined in the department of “Membrane Biochemistry” for Halobacterium salinarum, Natronomonas pharaonis, and Haloquadratum walsbyi, our group is analyzing all relevant classes of molecules by mass spectrometry. Currently the identification and quantitation of proteins, peptides, lipids, and small metabolites is performed. Connections between experimental and genome data are established database-driven.

A special emphasis of our work is on proteomics. In doing so, determination of the protein inventory helps to overcome the difficulties in gene prediction of GC-rich genomes. The successful development of methods for the analysis of “difficult” sub-proteoms like the “membrane proteome” and the so-called “small proteome” (proteins below 25kD) enables the identification of more than 70% of all proteins predicted from the genome. The high coverage does not only allow for the unambiguous identification of many N-terminal peptides for the validation of the assigned start codons, but contributed also to the elucidation of N-terminal protein processing in halophilic archaea.

For the analysis of archaeal signal transduction networks post-translational modifications (phosphorylation, transducer methylation) are determined and complemented by absolute protein quantitation (copy number counting) and interaction studies (EU project INTERACTOME). By means of quantitative proteome experiments (ICPL, SILAC) relative amounts of protein species are measured in response to different growth conditions of the cell. Informations about the archaeal metabolism collected by this procedure can subsequently be improved by a direct GC/MS analysis of selected metabolites. As an example, one project is about the elucidation of details from the isoprenoid metabolism by the use of stable isotope labeling in GC/MS based lipid analysis.

All data collected by our group are integrated into the database system “HaloLex”, which was developed by the project group “Bioinformatics” in our department. This system provides access to experimental data for theoretical analyses in the context of system biology of halophilic archaea.