Project number: Z1-70013
Period: 1.3.2026 – 29.2.2028
Head: dr. Gašper Šolinc
Cryo-electron microscopy (cryo-EM) is a key technique for determining the atomic structures of membrane proteins (MPs), which represent 27% of the human proteome. Despite the success of this technique, challenges remain, such as the poor solubility of MPs, their heterogeneity, low signal, and the difficulty of providing a native lipid environment.
To overcome these hurdles, as part of this postdoctoral project, we are developing a method for forming planar lipid bilayers (PLBs) directly on EM grids. Compared to the use of detergents or small lipid vesicles, PLBs provide a native environment with low curvature, which improves the signal-to-noise ratio and enables the observation of larger protein complexes. We plan to fabricate the PLBs by passing the grid through a non-polar lipid solution into a polar phase, where the lipid monolayer spontaneously transforms into a stable bilayer.
Main Advantages of the Approach:
- High Data Quality: The homogeneous distribution of the bilayer over a large grid surface area allows for the collection of a greater volume of data.
- Dynamic Studies: Since the PLB is present on the grid prior to the addition of proteins, we can precisely control the interaction time and capture various stages of dynamic processes via vitrification.
- Biological Relevance: The structure of a PLB is closer to real plasma membranes than highly curved vesicles.
The method will be validated by studying pore-forming proteins (PFPs), such as listeriolysin O (LLO), Fav, and equinatoxin II. Upon binding to receptors, these proteins oligomerize into complex structures, representing an ideal model for testing the spatial and temporal resolution of this new cryo-EM approach.