We usually have openings for enthusiastic and hard-working Masters students in our group, depending upon the availability of lab members who are able to supervise students. We have hosted students pursuing their Bachelor and Masters End Projects from Nanobiology, Life Sciences and Technology, and Applied Physics programs. Bachelors and Masters students have been co-authors (and first authors) on several of our publications, including our work published in Nature Chemical Biology, eLife, and Nature Communications.
Currently, BEP/MEP projects will be focused on membrane metabolism. Contact g.e.bokinsky ((at)) tudelft.nl for more information.
Metabolic regulation of bacterial fatty acid biosynthesis. Every cell has a membrane. Membranes need to be kept intact, have the right composition, and constantly adjust to the environment. This is challenging to study because these responses are not driven by transcriptional regulation, but by proteins and enzymes sensing conditions and directing responses themselves. This form of regulation (called "post-translational regulation") is analogous to our autonomous nervous system, which manages our heartbeat, breathing rate, and digestion so our conscious minds don’t have to. Since the membrane is the most basic foundation of every cell envelope, our findings are highly relevant to understanding how antibiotics kill bacteria.
Our projects use both live-cell fluorescence microscopy and mass spectrometry -- two cutting-edge techniques that generate cool data. We also use tools from synthetic biology to engineer bacteria with interesting and useful properties. Our mass spectrometry specialty is in quantifying the enzymes and metabolic intermediates of membrane synthesis. In other words, we can see the entire membrane assembly line, which gives us information about which steps are controlled to produce the interesting behaviours we study.
A membrane protein of unknown function has recently been found to control fatty acid synthesis. We have the tools to discover how this happens. The interested student will compare the membrane synthesis pathway of strains lacking this unusual protein to determine which step in the fatty acid or phospholipid pathways is regulated by this protein. If time allows, the student will also construct a fluorescently-tagged version of the protein to follow its localization in different growth and membrane stress conditions.
Different lipids exhibit different preferences for membrane shapes. For instance, cardiolipin is often found in highly-curved membranes, such as the poles of bacteria and the inner cristae of mitochondria. We have preliminary evidence that the enzyme that generates cardiolipin may be directly activated by increased membrane curvature, and that the reverse reaction (breaking cardiolipin back into two phospholipids) is also driven by decreased membrane curvature. The interested student will quantify cardiolipin and other phospholipids in E. coli cells waking up from dormancy and in cells engineered to generate small and highly-curved minicells.
The outer membrane of Gram-negative bacteria is asymmetric: the inner leaflet is the "usual" phospholipid leaflet, while the outer leaflet is essential steel-plate armor (lipopolysaccharides). Lipopolysaccharides are why Gram-negative bacteria are hard to kill. However, bacteria must precisely balance phospholipid production (which form "soft" membranes) with lipopolysaccharide production ("hard" membranes)… or die. (No joke.) The balancing mechanism is highly controversial, but it is thought to depend upon protein-protein interactions that detect the amount of lipopolysaccharides accumulated along the inner membrane waiting to be transported to the outer membrane. The student will construct a fluorescent biosensor that will report the protein-protein interactions in real time and determine whether this interaction is sufficient to balance phospholipid production with lipopolysaccharide production.