Host-pathogen interactions:
mammalian-bacterial, honeybee-bacterial, honeybee-mite.
Our lab is focused on quantitative proteomics using LC-ESI-MS to study biological systems. We cover a wide range of topics including pathogen invasion, infection, protein localization, and mapping protein interaction.
Mass spectrometry-based proteomics:
In order to understand how a complex system works it is extremely useful to know the identities of as many of the components of the system as possible. Commercially available mass spectrometers and database search tools have advanced to the point where experienced laboratories can routinely and reliably identify hundreds or thousands of proteins in exceedingly complex mixtures (i.e. an organelle preparation) using nanoflow high performance liquid chromatography (HPLC)/tandem mass spectrometry (LC/MS). Typically, complex protein samples are reduced to peptides by highly specific enzymatic digestion (e.g. trypsin cleavage carboxy-terminal of Arg or Lys). The peptides are then resolved by extremely high-resolution reversed-phase chromatography and eluted/ionized directly into a mass spectrometer (MS). The MS, operating in an information-dependent mode, selects and isolates any peptide ions observed and subjects them to tandem mass spectrometry. The observed mass/charge ratio of the peptide and its fragmentation pattern are then used to scan comprehensive protein sequence libraries (e.g. for a human liver sample the data would be searched against the Homo sapiens library) to find the best theoretical match and the peptides are compiled to arrive at a ΓÇÿprotein hit list'.
In the past few years we have witnessed great advances in methodologies for applying mass spectrometry-based proteomics to answer biologically relevant questions. Some of the most powerful of these methods involve the use of stable isotopes to incorporate a quantitative dimension into the experiment. Thus far, however, quantitative proteomics has yet to be widely applied in mainstream biology. Our group is interested in a multidisciplinary approach to relevant cell biological problems. We develop and apply quantitative proteomic methods to questions involving organelles and how the composition of these compartments change when antagonized. Additionally, we are involved in several projects involving host-pathogen interactions. The knowledge gained from proteomic experiments is then used to direct more in-depth biochemical, bioinformatic and cell biological analysis of the system in order to validate the proteomic results. We have an LTQ-Orbitrap and an LTQ-FT in our laboratory, both equipped with autosamplers and nESI-interfaced HPLCs.
