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Title: Purification, crystallization and preliminary crystallographic analysis of a GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus

Abstract

A GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus has been crystallized. Combined with biochemical analyses, it is expected that the structure of this protein will give insight in the function of a relatively unknown subfamily of the GTPase superfamily. A predicted GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus, termed SsGBP, has been cloned and overexpressed in Escherichia coli. The purified protein was crystallized using the hanging-drop vapour-diffusion technique in the presence of 0.05 M cadmium sulfate and 0.8 M sodium acetate pH 7.5. A single-wavelength anomalous dispersion data set was collected to a maximum resolution of 2.0 Å using a single cadmium-incorporated crystal. The crystal form belongs to space group P2{sub 1}2{sub 1}2{sub 1}, with approximate unit-cell parameters a = 65.0, b = 72.6, c = 95.9 Å and with a monomer in the asymmetric unit.

Authors:
;  [1];  [2];  [1];  [3];  [1];  [3];  [1]
  1. Laboratory of Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Hesselink van Suchtelenweg 4, 6703 CT Wageningen (Netherlands)
  2. (China)
  3. National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 (China)
Publication Date:
OSTI Identifier:
22360274
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section F; Journal Volume: 63; Journal Issue: Pt 3; Other Information: PMCID: PMC2330175; PMID: 17329823; PUBLISHER-ID: bw5186; OAI: oai:pubmedcentral.nih.gov:2330175; Copyright (c) International Union of Crystallography 2007; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United Kingdom
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; ACETATES; CADMIUM; CRYSTALLIZATION; CRYSTALS; DIFFUSION; DISPERSIONS; ESCHERICHIA COLI; MONOMERS; PROTEINS; RESOLUTION; SODIUM; SPACE GROUPS; WAVELENGTHS

Citation Formats

Wu, Hao, Sun, Lei, National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, Brouns, Stan J. J., Fu, Sheng, Akerboom, Jasper, Li, Xuemei, E-mail: lixm@sun5.ibp.ac.cn, and Oost, John van der, E-mail: lixm@sun5.ibp.ac.cn. Purification, crystallization and preliminary crystallographic analysis of a GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus. United Kingdom: N. p., 2007. Web. doi:10.1107/S1744309107008500.
Wu, Hao, Sun, Lei, National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, Brouns, Stan J. J., Fu, Sheng, Akerboom, Jasper, Li, Xuemei, E-mail: lixm@sun5.ibp.ac.cn, & Oost, John van der, E-mail: lixm@sun5.ibp.ac.cn. Purification, crystallization and preliminary crystallographic analysis of a GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus. United Kingdom. doi:10.1107/S1744309107008500.
Wu, Hao, Sun, Lei, National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, Brouns, Stan J. J., Fu, Sheng, Akerboom, Jasper, Li, Xuemei, E-mail: lixm@sun5.ibp.ac.cn, and Oost, John van der, E-mail: lixm@sun5.ibp.ac.cn. Thu . "Purification, crystallization and preliminary crystallographic analysis of a GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus". United Kingdom. doi:10.1107/S1744309107008500.
@article{osti_22360274,
title = {Purification, crystallization and preliminary crystallographic analysis of a GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus},
author = {Wu, Hao and Sun, Lei and National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 and Brouns, Stan J. J. and Fu, Sheng and Akerboom, Jasper and Li, Xuemei, E-mail: lixm@sun5.ibp.ac.cn and Oost, John van der, E-mail: lixm@sun5.ibp.ac.cn},
abstractNote = {A GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus has been crystallized. Combined with biochemical analyses, it is expected that the structure of this protein will give insight in the function of a relatively unknown subfamily of the GTPase superfamily. A predicted GTP-binding protein from the hyperthermophilic archaeon Sulfolobus solfataricus, termed SsGBP, has been cloned and overexpressed in Escherichia coli. The purified protein was crystallized using the hanging-drop vapour-diffusion technique in the presence of 0.05 M cadmium sulfate and 0.8 M sodium acetate pH 7.5. A single-wavelength anomalous dispersion data set was collected to a maximum resolution of 2.0 Å using a single cadmium-incorporated crystal. The crystal form belongs to space group P2{sub 1}2{sub 1}2{sub 1}, with approximate unit-cell parameters a = 65.0, b = 72.6, c = 95.9 Å and with a monomer in the asymmetric unit.},
doi = {10.1107/S1744309107008500},
journal = {Acta Crystallographica. Section F},
number = Pt 3,
volume = 63,
place = {United Kingdom},
year = {Thu Mar 01 00:00:00 EST 2007},
month = {Thu Mar 01 00:00:00 EST 2007}
}
  • Type 1 RNase H from the hyperthermophilic archaeon S. tokodaii 7 was overproduced in E. coli, purified, and crystallized. Preliminary crystallographic studies indicated that the crystal belongs to space group P4{sub 3}, with unit-cell parameters a = b = 39.21, c = 91.15 Å. Crystallization and preliminary crystallographic studies of type 1 RNase H from the hyperthermophilic archaeon Sulfolobus tokodaii 7 were performed. A crystal was grown at 277 K by the sitting-drop vapour-diffusion method. Native X-ray diffraction data were collected to 1.5 Å resolution using synchrotron radiation from station BL41XU at SPring-8. The crystal belongs to space group P4{submore » 3}, with unit-cell parameters a = b = 39.21, c = 91.15 Å. Assuming the presence of one molecule in the asymmetric unit, the Matthews coefficient V{sub M} was calculated to be 2.1 Å{sup 3} Da{sup −1} and the solvent content was 40.5%. The structure of a selenomethionine Sto-RNase HI mutant obtained using a MAD data set is currently being analysed.« less
  • The C-terminal domain of the transcriptional regulator Ss-LrpB from S. solfataricus was purified by affinity chromatography and crystallized. Crystals belong to space group P2{sub 1}2{sub 1}2. A complete data set was collected to a resolution of 2 Å. Ss-LrpB from Sulfolobus solfataricus P2 belongs to the bacterial/archaeal superfamily of Lrp-like (leucine-responsive regulatory protein-like) transcription regulators. The N-terminal DNA-binding domain contains a HTH motif and the C-terminal domain contains an αβ-sandwich (βαββαβ fold). The C-terminal domain was overexpressed in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. The crystals belong to space group P2{sub 1}2{sub 1}2, with unit-cell parametersmore » a = 59.35, b = 74.86, c = 38.08 Å and a data set was collected to 2.0 Å resolution. Structure determination using a selenomethionine derivative is under way.« less
  • A phosphotriesterase (PTE) from the hyperthermophilic archaeon S. solfataricus has been crystallized. Combined with biochemical and bioengineering studies, it is expected that the structure of this protein will provide insight into the natural function of the PTE family and provide important data for achieving an efficient organophosphate biodecontaminant. Organophosphates constitute the largest class of insecticides used worldwide and some of them are potent nerve agents. Consequently, organophosphate-degrading enzymes are of paramount interest as they could be used as bioscavengers and biodecontaminants. Phosphotriesterases (PTEs) are capable of hydrolyzing these toxic compounds with high efficiency. A distant and hyperthermophilic representative of themore » PTE family was cloned from the archeon Sulfolobus solfataricus MT4, overexpressed in Escherichia coli and crystallized; the crystals diffracted to 2.54 Å resolution. Owing to its exceptional thermostability, this PTE may be an excellent candidate for obtaining an efficient organophosphate biodecontaminant. Here, the crystallization conditions and data collection for the hyperthermophilic S. solfataricus PTE are reported.« less
  • Nucleotide-exchange factor from S. solfataricus (SsEF-1β) has been successfully crystallized. X-ray diffraction data have been collected from the native enzyme and from the selenomethionine derivative of SsEF-1β to 1.97 and 1.83 Å resolution, respectively. The nucleotide-exchange factor isolated from the hyperthermophilic archaeon Sulfolobus solfataricus (SsEF-1β) consists of 90 residues and differs from eukaryal EF-1βs. The protein has been successfully crystallized using either microbatch-under-oil or vapour-diffusion methods. Crystals of native SsEF-1β diffract to 1.97 Å resolution and belong to space group P2{sub 1}2{sub 1}2, with unit-cell parameters a = 106.46, b = 54.87, c = 44.03 Å. Diffraction data have alsomore » been collected from a selenomethionine derivative of SsEF-1β at 1.83 Å resolution. Model building using the phases derived from the MAD experiment is in progress.« less
  • The thioredoxin reductase isolated from S. solfataricus has been crystallized. Diffraction data have been collected from the wild type and from the NADP-bound form of the enzyme to 1.80 and 1.95 Å, respectively. The structure of the thioredoxin reductase has been solved using MAD diffraction data. A thermostable thioredoxin reductase isolated from Sulfolobus solfataricus (SsTrxR) has been successfully crystallized in the absence and in the presence of NADP. Two different crystal forms have been obtained. Crystals of the form that yields higher resolution data (1.8 Å) belong to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 76.77,more » b = 120.68, c = 126.85 Å. The structure of the enzyme has been solved by MAD methods using the anomalous signal from the Se atoms of selenomethionine-labelled SsTrxR.« less