My major interests are in three-dimensional organization of large macromolecular complexes and cell organelles by electron microscopy, cryo-electron microscopy (cryoEM) in particular. CryoEM allows to study symmetric particles, e.g. spherical viruses, ordered assemblies (2D crystals, helical arrays, etc.) and asymmetrical complexes in their native state. Unique structures are studied using electron tomography. I am interested in structural studies of symmetrical as well as of asymmetrical particles by these techniques.
Spherical viruses are highly symmetrical; they infect variety of animal and plant cells. The structure opens way to understand their mechanism of action and allows to target specific epitopes to neutralize them. As an example, in Red clover necrotic mosaic virus (RCNMV), whose structure we determined at 8Å resolution, substantial portion of its genome is ordered and forms a distinct cage within the viral capsid. We found that depletion of Ca and Mg from RCNMV causes structural changes resulting in opening of ~13 Å diameter channels at certain locations in the capsid allowing RNA to egress and start replication. Recently determined structure of Western equine encephalitis virus is the first example of a wild type BSL-3 agent spatial organization determined by cryoEM with imaging performed in a BSL-3 containment, a unique laboratory space available at UTMB. In addition to traditional cryoEM of single particles we used cryo-electron tomography to study three-dimensional organization of a bunyavirus Rift Valley fever virus which at the time was unknown to have any symmetry. We plan now to apply that method to study cells, cellular organelles and cell-pathogen interactions.
Asymmetrical assemblies are the most challenging case for structural studies. An example is LDL, one of the major lipid carriers in blood. They have only one protein component, apolipoprotein B-100, whereas the rest of the particle (78%) is composed of various lipids. Amazingly, despite their high lipid content, they are rigid enough to maintain their overall shape and characteristic lamellar organization of the core. We found that the lamellae most likely consist of cholesteryl esters; therefore with low or no cholesterol in LDL they do not have striated cores and are less rigid. The findings provide an important hypothesis that LDL depending on cholesterol content would interact with LDL receptors differently causing lipid metabolic disorders.