Journal of Structural Biology 151 (2005) 111–115 www.elsevier.com/locate/yjsbi 1047-8477/$ - see front matter  2005 Elsevier Inc. All rights reserved. doi:10.1016/j.jsb.2005.04.002 Crystallization note Preliminary analysis of two and three dimensional crystals of vault ribonucleoprotein particles Jordi Querol-Audí a , Rosa Perez-Luque a , Ignacio Fita a , Carmen Lopéz-Iglesias b , José R. Castón c , José L. Carrascosa c , Nuria Verdaguer a,¤ a Instituto de Biología Molecular de Barcelona (CSIC-Parc CientíWc), Josep-Samitier 1-5, Barcelona 08028, Spain b Unitat de Microscòpia Electrònica (SCT-Universitat de Barcelona, Parc Científíc), Josep-Samitier 1-5, Barcelona 08028, Spain c Centro Nacional de Biotecnología/CSIC, Campus Universidad Autónoma de Madrid, 28049 Madrid, Spain Received 21 February 2005, received in revised form 13 April 2005, and accepted 15 April 2005 Available online 10 May 2005 Abstract Vaults are large ribonucleoprotein particles found in a wide variety of eukaryotes. When imaged by electron-microscopy vaults present a strikingly conserved barrel-shaped structure with an invaginated waist and two protruding caps. In this work, we present two dimensional (2D) and three dimensional (3D) crystals of naturally produced vaults in murine and monkey cells, respectively. The 2D-crystals presented a hexagonal packing with the lattice parameter deWned by the diameter of the vault barrel. Fourier transforms from images of the negatively stained 2D-crystals showed spots till about 45 Å resolution. The 3D-crystals reached about 0.15 £ 0.15 £ 0.02 mm 3 in size and presented a Xat triangular morphology with well-developed faces. The preliminary characteriza- tion of these 3D-crystals, which diVract very weakly to »10 Å resolution, suggests a trigonal packing with the R32 space group symmetry. The 3D-crystals appear to be formed by adding layers of vaults, which retain the hexagonal organization seen in the 2D-crystals, with relative shifts that maximize the interdigitation of particles in adjacent layers. Accurate crystal symmetry in the 2D- and 3D-crystals requires neighbor particles interacting according to a 6-fold and a 3-fold dihedral symmetry, respectively. Compatibility with the reported 8-fold symmetry would imply multiples of 24-fold rotational symmetry, in agreement with the recently proposed 48-fold dihedral symmetry for reconstituted recombinant vaults.  2005 Elsevier Inc. All rights reserved. Keywords: Vault; Ribonucleoprotein; Electron microscopy; X-ray crystallography 1. Introduction With a mass of 13-MDa and overall dimensions of 40 £ 70 nm, the vault complex is the largest ribonucleo- protein particle found in eukaryotes as diverse as mam- mals, avians, amphibians, and the lower eukaryote Dictyostelium discoideum (Hamill and Suprenant, 1997; Herrmann et al., 1998; Kedersha et al., 1990). In mam- mals, vaults contain three major proteins: the 100 kDa major vault protein (MVP; Kedersha and Rome, 1986), the 193 kDa vault poly(ADP-ribosyl)ating polymerase (VPARP; Kickhoefer et al., 1999a), and the 240 kDa tel- omerase-associated protein (TEP1; Kickhoefer et al., 1999b). Additionally, one small untranslated RNA at least is found as a constitutive component (Kickhoefer et al., 1993). Despite their diverse origin, vaults are uniform in size and morphology when imaged by electron-microscopy presenting a barrel-like structure with an invaginated waist and two protruding caps (Kong et al., 1999). Approximately 75% of the vault particle is due to the MVP (Kedersha et al., 1991). Vault-like particles, similar to puriWed endogenous vaults, are observed when the rat MVP is expressed in insect cells (Stephen et al., 2001). * Corresponding author. Fax: +34 93 4034979. E-mail address: nvmcri@ibmb.csic.es (N. Verdaguer).