Browsing by Author "Vlassakis, Julea"

Now showing 1 - 2 of 2
  • Thumbnail Image
    ItemEmbargo
    Embedding Porous Polymers Films in Grids Towards Resolving Intrinsically Disordered Proteins with Cryo-EM Imaging
    (2024-04-19) Leung, Wesley; Vlassakis, Julea
    Ewing’s sarcoma is a rare cancer that predominantly occurs in young adults and children. 80-90% of Ewing’s sarcoma cases contain an oncogenic protein EWS/FLI1. While it is known to be an intrinsically disordered protein (IDP), little is known about the protein itself, including its structure. To reveal its structure through cryo-electron microscopy (cryo-EM), we attempt to develop a method in the sample preparation stage to reveal the structure of EWS/FLI1 and other IDPs. Support films have been commonly used to increase the throughput of the sample distribution and other functionalities. We aim to add a continuous hydrogel film for the IDPs to bind and stabilize for screening. With the simulation of EWS/FLI1 folding, we revealed the surface properties of EWS/FLI1 and characterized our continuous film composed of poly(2-hydroxyethyl methacrylate) (pHEMA). We also showed negative staining of thrombin in preparation to show that our pHEMA film can assist in resolving IDPs.
  • Thumbnail Image
    Item
    Microscale measurements of protein complexes from single cells
    (Elsevier, 2024) Dutta, Tanushree; Vlassakis, Julea
    Proteins execute numerous cell functions in concert with one another in protein–protein interactions (PPI). While essential in each cell, such interactions are not identical from cell to cell. Instead, PPI heterogeneity contributes to cellular phenotypic heterogeneity in health and diseases such as cancer. Understanding cellular phenotypic heterogeneity thus requires measurements of properties of PPIs such as abundance, stoichiometry, and kinetics at the single-cell level. Here, we review recent, exciting progress in single-cell PPI measurements. Novel technology in this area is enabled by microscale and microfluidic approaches that control analyte concentration in timescales needed to outpace PPI disassembly kinetics. We describe microscale innovations, needed technical capabilities, and methods poised to be adapted for single-cell analysis in the near future.