Dataset Information

Crystallization and preliminary X-ray crystallographic analysis of the NmrA-like DDB_G0286605 protein from the social amoeba Dictyostelium discoideum.

ABSTRACT: The DDB_G0286605 gene product from Dictyostelium discoideum, an NmrA-like protein that belongs to the short-chain dehydrogenase/reductase family, has been crystallized by the hanging-drop vapour-diffusion method at 295?K. A 1.64?Å resolution data set was collected using synchrotron radiation. The DDB_G0286605 protein crystals belonged to space group P2(1), with unit-cell parameters a=67.598, b=54.935, c=84.219?Å, ? = 109.620°. Assuming the presence of two molecules in the asymmetric unit, the solvent content was estimated to be about 43.25% with 99% probability. Molecular-replacement trials were attempted with three NmrA-like proteins, NmrA, HSCARG and QOR2, as search models, but failed. This may be a consequence of the low sequence identity between the DDB_G0286605 protein and the search models (DDB_G0286605 has a primary-sequence identity of 28, 32 and 19% to NmrA, HCARG and QOR2, respectively).

SUBMITTER: Kim MK

PROVIDER: S-EPMC3079982 | biostudies-literature | 2011 Jan

REPOSITORIES: biostudies-literature

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Crystallization and preliminary X-ray crystallographic analysis of the NmrA-like DDB_G0286605 protein from the social amoeba Dictyostelium discoideum.

Kim Min-Kyu MK Yim Hyung-Soon HS Kang Sa-Ouk SO

Acta crystallographica. Section F, Structural biology and crystallization communications 20101223 Pt 1

The DDB_G0286605 gene product from Dictyostelium discoideum, an NmrA-like protein that belongs to the short-chain dehydrogenase/reductase family, has been crystallized by the hanging-drop vapour-diffusion method at 295 K. A 1.64 Å resolution data set was collected using synchrotron radiation. The DDB_G0286605 protein crystals belonged to space group P2(1), with unit-cell parameters a=67.598, b=54.935, c=84.219 Å, β = 109.620°. Assuming the presence of two molecules in the asymmetric unit, the so ...[more]

PMID: 21206034

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Project description:Huntingtin is a large HEAT repeat protein first identified in humans, where a polyglutamine tract expansion near the amino terminus causes a gain-of-function mechanism that leads to selective neuronal loss in Huntington's disease (HD). Genetic evidence in humans and knock-in mouse models suggests that this gain-of-function involves an increase or deregulation of some aspect of huntingtin's normal function(s), which remains poorly understood. As huntingtin shows evolutionary conservation, a powerful approach to discovering its normal biochemical role(s) is to study the effects caused by its deficiency in a model organism with a short life-cycle that comprises both cellular and multicellular developmental stages. To facilitate studies aimed at detailed knowledge of huntingtin's normal function(s), we generated a null mutant of hd, the HD ortholog in Dictyostelium discoideum. Dictyostelium cells lacking endogenous huntingtin were viable but during development did not exhibit the typical polarized morphology of Dictyostelium cells, streamed poorly to form aggregates by accretion rather than chemotaxis, showed disorganized F-actin staining, exhibited extreme sensitivity to hypoosmotic stress, and failed to form EDTA-resistant cell-cell contacts. Surprisingly, chemotactic streaming could be rescued in the presence of the bivalent cations Ca(2+) or Mg(2+) but not pulses of cAMP. Although hd(-) cells completed development, it was delayed and proceeded asynchronously, producing small fruiting bodies with round, defective spores that germinated spontaneously within a glassy sorus. When developed as chimeras with wild-type cells, hd(-) cells failed to populate the pre-spore region of the slug. In Dictyostelium, huntingtin deficiency is compatible with survival of the organism but renders cells sensitive to low osmolarity, which produces pleiotropic cell autonomous defects that affect cAMP signaling and as a consequence development. Thus, Dictyostelium provides a novel haploid organism model for genetic, cell biological, and biochemical studies to delineate the functions of the HD protein.

Dataset Information

Crystallization and preliminary X-ray crystallographic analysis of the NmrA-like DDB_G0286605 protein from the social amoeba Dictyostelium discoideum.

Publications

Crystallization and preliminary X-ray crystallographic analysis of the NmrA-like DDB_G0286605 protein from the social amoeba Dictyostelium discoideum.

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