Biophysical Society Thematic Meeting | Trieste 2024

Emerging Theoretical Approaches to Complement Single-Particle Cryo-EM

Poster Abstracts

13-POS Board 13 CRYO-EM STRUCTURE OF THE 8 MDA HUMAN VAULT PARTICLE AT 3.2 ÅNGSTROM Fabio Lapenta 1,2 ; Jana Aupic 3 ; Simone Marrancone 2 ; Sonia Covaceuszach 6 ; Gangupam Bhavani 2 ; Alexandre Durand 4 ; Nils Marechal 4 ; Dihia Moussaoui 7 ; Claudia d'Ercole 2 ; De March Matteo 2 ; Ario de Marco 2 ; Davide Cotugno 5 ; Gianni Frascotti 5 ; Paolo Tortora 5 ; Alessandra Magistrato 3 ; Alberto Cassetta 6 ; 1 International Center for Genetic Engineering and Biotechnology, Tumour virology, Trieste, Italy 2 University of Nova Gorica, Laboratory for Environmental and Life Sciences , Nova Gorica, Slovenia 3 National Research Council of Italy , Materials Foundry, Trieste, Italy 4 Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch , France 5 Department of Biotechnology and Biosciences, University of Milano-Bicocca, Milano, Italy 6 National Research Council of Italy, Institut of Crystallography, Trieste, Italy 7 European Synchrotron Radiation Facility, Grenoble, France Vaults are large ribonucleoprotein (RNP) particles with a size of 70 nm × 40 nm × 40 nm. Initially reported in 1986, vaults have a distinct ovoidal architecture made of 2 symmetric 39 meric shells composed of the major vault protein (MVP). Despite its large size and the high levels of conservation of the vault components, the precise role of this complex in the cell is not yet fully understood. Here, we report the first cryoelectron microscopy (cryoEM) reconstruction of the human vault complex at 3.2 Å obtained with a standard workflow in cryoSPARC and provide a comparison to the previously published structures of its murine counterpart. Additionally, we investigated the interaction and binding site to the human onco-suppressor protein PTEN with Grating-Coupled Interferometry (GCI) and Small Angle X-ray scattering (SAXS), which showed a strong affinity (kd of 1 µM) in presence of 10 mM of Ca(II), this supports the role of the vault as molecular scaffold and transport of the onco-suppressor protein.

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