title: Localization and orientation of heavy-atom cluster compounds in protein crystals using molecular replacement.
abstract: Heavy-atom clusters (HA clusters) containing a large number of specifically arranged electron-dense scatterers are especially useful for experimental phase determination of large complex structures, weakly diffracting crystals or structures with large unit cells. Often, the determination of the exact orientation of the HA cluster and hence of the individual heavy-atom positions proves to be the critical step in successful phasing and subsequent structure solution. Here, it is demonstrated that molecular replacement (MR) with either anomalous or isomorphous differences is a useful strategy for the correct placement of HA cluster compounds. The polyoxometallate cluster hexasodium α-metatungstate (HMT) was applied in phasing the structure of death receptor 6. Even though the HA cluster is bound in alternate partially occupied orientations and is located at a special position, its correct localization and orientation could be determined at resolutions as low as 4.9 Å. The broad applicability of this approach was demonstrated for five different derivative crystals that included the compounds tantalum tetradecabromide and trisodium phosphotungstate in addition to HMT. The correct placement of the HA cluster depends on the length of the intramolecular vectors chosen for MR, such that both a larger cluster size and the optimal choice of the wavelength used for anomalous data collection strongly affect the outcome.
authors: Protein Crystallography Group, Leibniz Institute for Age Research - Fritz Lipmann Institute (FLI), Beutenbergstrasse 11, D-07745 Jena, Germany. sdahms@fli-leibniz.de
authors: Sven O
authors: SO
authors: Dahms
authors: Miriam
authors: M
authors: Kuester
authors: Carsten
authors: C
authors: Streb
authors: Christian
authors: C
authors: Roth
authors: Norbert
authors: N
authors: Sträter
authors: Manuel E
authors: ME
authors: Than
mesh_terms: D000818:Animals; D018360:Crystallography, X-Ray / Q000379:methods* / Q000592:standards; D030562:Databases, Protein / Q000592:standards; D004583:Electrons; D006801:Humans; D019216:Metals, Heavy / Q000737:chemistry; D051379:Mice; D008958:Models, Molecular; D008970:Molecular Weight; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism; D018124:Receptors, Tumor Necrosis Factor / Q000737:chemistry* / Q000378:metabolism; D012542:Scattering, Radiation; D044764:Ubiquitin-Activating Enzymes / Q000737:chemistry* / Q000378:metabolism
chemical_list: D019216:Metals, Heavy; D018124:Receptors, Tumor Necrosis Factor; C114109:TNFRSF21 protein, human; D009113:Muramidase; D044764:Ubiquitin-Activating Enzymes
keywords: death receptor 6; experimental phasing; heavy-metal cluster; hexasodium α-metatungstate; molecular replacement
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