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Title: Complex assembly, crystallization and preliminary X-ray crystallographic analysis of the human Rod–Zwilch–ZW10 (RZZ) complex

The 800 kDa complex of the human Rod, Zwilch and ZW10 proteins (the RZZ complex) was reconstituted in insect cells, purified, crystallized and subjected to preliminary X-ray diffraction analysis. The spindle-assembly checkpoint (SAC) monitors kinetochore–microtubule attachment during mitosis. In metazoans, the three-subunit Rod–Zwilch–ZW10 (RZZ) complex is a crucial SAC component that interacts with additional SAC-activating and SAC-silencing components, including the Mad1–Mad2 complex and cytoplasmic dynein. The RZZ complex contains two copies of each subunit and has a predicted molecular mass of ∼800 kDa. Given the low abundance of the RZZ complex in natural sources, its recombinant reconstitution was attempted by co-expression of its subunits in insect cells. The RZZ complex was purified to homogeneity and subjected to systematic crystallization attempts. Initial crystals containing the entire RZZ complex were obtained using the sitting-drop method and were subjected to optimization to improve the diffraction resolution limit. The crystals belonged to space group P3{sub 1} (No. 144) or P3{sub 2} (No. 145), with unit-cell parameters a = b = 215.45, c = 458.7 Å, α = β = 90.0, γ = 120.0°.
Authors:
;  [1] ;  [2] ;  [1] ;  [1] ;  [3]
  1. Max Planck Institute of Molecular Physiology, Otto Hahn Strasse 11, 44227 Dortmund (Germany)
  2. Institut Curie, CNRS UMR 3348/INSERM U1005, Bâtiment 110, Centre Universitaire, 91405 Orsay CEDEX (France)
  3. (Germany)
Publication Date:
OSTI Identifier:
22375717
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta crystallographica. Section F, Structural biology communications; Journal Volume: 71; Journal Issue: Pt 4; Other Information: PMCID: PMC4388180; PMID: 25849506; PUBLISHER-ID: pg5052; PUBLISHER-ID: S2053230X15004343; OAI: oai:pubmedcentral.nih.gov:4388180; Copyright (c) Altenfeld et al. 2015; This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CRYSTALLIZATION; CRYSTALS; MASS; MICROTUBULES; OPTIMIZATION; PROTEINS; RESOLUTION; RODS; SPACE GROUPS; SPIN; X-RAY DIFFRACTION