Rodriguez de Francisco, B.

bioRxiv2022

MEPSi: A tool for simulating tomograms of membrane-embedded proteins

The throughput and fidelity of cryogenic cellular electron tomography (cryo-ET) is constantly increasing through advances in cryogenic electron microscope hardware, direct electron detection devices, and powerful image processing algorithms. However, the need for careful optimization of sample preparations and for access to expensive, high-end equipment, make cryo-ET a costly and time-consuming technique. Generally, only after the last step of the cryo-ET workflow, when reconstructed tomograms are available, it becomes clear whether the chosen imaging parameters were suitable for a specific type of sample in order to answer a specific biological question. Tools for a-priory assessment of the feasibility of samples to answer biological questions and how to optimize imaging parameters to do so would be a major advantage. Here we describe MEPSi (Membrane Embedded Protein Simulator), a simulation tool aimed at rapid and convenient evaluation and optimization of cryo-ET data acquisition parameters for studies of transmembrane proteins in their native environment. We demonstrate the utility of MEPSi by showing how to detangle the influence of different data collection parameters and different orientations in respect to tilt axis and electron beam for two examples: (1) simulated plasma membranes with embedded single-pass transmembrane IIb{beta}3 integrin receptors and (2) simulated virus membranes with embedded SARS-CoV-2 spike proteins. HIGHLIGHTSO_LITool to simulate tomograms of membrane-embedded proteins C_LIO_LIDetangles influence of data acquisition parameters from sample quality issues C_LIO_LIRapid evaluation and optimization of cryo-ET data acquisition parameters C_LIO_LIProof-of-concept provided with integrins and SARS-CoV-2 spike simulations C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=132 SRC="FIGDIR/small/501771v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@120b44eorg.highwire.dtl.DTLVardef@1d9056org.highwire.dtl.DTLVardef@ef327aorg.highwire.dtl.DTLVardef@1daf773_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2022

ACE2 nanoparticles prevent cell entry of SARS-CoV-2

The continual evolution of SARS-CoV-2 has challenged the efficacy of many COVID19 vaccines and treatment options. One strategy that evades viral escape is using the entry receptor, human Angiotensin-Converting Enzyme 2 (hACE2). Soluble hACE2 receptor domains show potential as decoys but genetic modifications are necessary to provide sufficient efficacy. However, these engineered constructs are potentially susceptible to viral escape. We combined native hACE2 with viral vectors to form nanoparticles presenting hACE2 analogous to human cells. Cell-based viral infection assays and cryogenic in-situ tomography show that hACE2 nanoparticles sequester viruses through aggregation, efficiently blocking entry of SARS-CoV-2 and its variants in model cell systems and human respiratory tract explants using native hACE2. Thus, we show that hACE2 nanoparticles have high potential as pan-variant COVID19 therapeutics.

cell biology↗