Modeling the structure of pyrococcus furiosus rubredoxin by homology to other X-ray structures (original) (raw)
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Protein Science, 1992
The three-dimensional solution-state structure is reported for the zinc-substituted form of rubredoxin (Rd) from the marine hyperthermophilic archaebacterium Pyrococcus furiosus, an organism that grows optimally at 100 " c . Structures were generated with DSPACEB by a hybrid distance geometry (DG)-based simulated annealing (SA) approach that employed 403 nuclear Overhauser effect (N0E)-derived interproton distance restraints, including 67 interresidue, 124 sequential ( i -j = I), 75 medium-range ( i -j = 2-5), and 137 long-range ( i -j > 5) restraints.
Structure of the hypothetical protein PF0899 from Pyrococcus furiosus at 1.85 Å resolution
Acta Crystallographica Section F Structural Biology and Crystallization Communications, 2007
The hypothetical protein PF0899 is a 95-residue peptide from the hyperthermophilic archaeon Pyrococcus furiosus that represents a gene family with six members. P. furiosus ORF PF0899 has been cloned, expressed and crystallized and its structure has been determined by the Southeast Collaboratory for Structural Genomics (http://www.secsg.org). The structure was solved using the SCA2Structure pipeline from multiple data sets and has been refined to 1.85 Å against the highest resolution data set collected (a presumed gold derivative), with a crystallographic R factor of 21.0% and R free of 24.0%. The refined structure shows some structural similarity to a wedge-shaped domain observed in the structure of the major capsid protein from bacteriophage HK97, suggesting that PF0899 may be a structural protein.
Crystal structure of rubredoxin from Desulfovibrio gigas to ultra-high 0.68Å resolution
Biochemical and Biophysical Research Communications, 2006
Rubredoxin (D.g. Rd) is a small non-heme iron-sulfur protein shown to function as a redox coupling protein from the sulfate reducing bacteria Desulfovibrio gigas. The protein is generally purified from anaerobic bacteria in which it is thought to be involved in electron transfer or exchange processes. Rd transfers an electron to oxygen to form water as part of a unique electron transfer chain, composed by NADH:rubredoxin oxidoreductase (NRO), rubredoxin and rubredoxin:oxygen oxidoreductase (ROO) in D.g. The crystal structure of D.g. Rd has been determined by means of both a Fe single-wavelength anomalous dispersion (SAD) signal and the direct method, and refined to an ultra-high 0.68 Å resolution, using X-ray from a synchrotron. Rd contains one iron atom bound in a tetrahedral coordination by the sulfur atoms of four cysteinyl residues. Hydrophobic and p-p interactions maintain the internal Rd folding. Multiple conformations of the iron-sulfur cluster and amino acid residues are observed and indicate its unique mechanism of electron transfer. Several hydrogen bonds, including NAHÁ Á ÁSG of the iron-sulfur, are revealed clearly in maps of electron density. Abundant waters bound to CAO peptides of residues Val8, Cys9, Gly10, Ala38, and Gly43, which may be involved in electron transfer. This ultrahigh-resolution structure allows us to study in great detail the relationship between structure and function of rubredoxin, such as salt bridges, hydrogen bonds, water structures, cysteine ligands, iron-sulfur cluster, and distributions of electron density among activity sites. For the first time, this information will provide a clear role for this protein in a strict anaerobic bacterium.
Proceedings of the National Academy of Sciences, 2004
The structure of a partially deuterated rubredoxin from the hyperthermophilic archaeon Pyrococcus furiosus, an organism that grows optimally at 100°C, was determined by using the neutron single-crystal diffractometer dedicated for biological macromolecules (BIX-3) at the JRR-3M reactor of the Japan Atomic Energy Research Institute. Data were collected at room temperature up to a resolution of 1.5 Å, and the completeness factor of the data set was 81.9%. The model contains 306 H and 50 D atoms. A total of 37 hydration water molecules were identified, with 15 having all three atoms fully located and the remaining D 2O molecules partially defined. The model has been refined to final agreement factors of R ؍ 18.6% and R free ؍ 21.7%. Several orientations of the O-D bonds of side chains, whose assignments from x-ray data were previously ambiguous, were clearly visible in the neutron structure. Although most backbone N-H bonds had undergone some degree of H͞D exchange throughout the rubredoxin molecule, 5 H atom positions still had distinctly negative (H) peaks. The neutron Fourier maps clearly showed the details of an extensive set of H bonds involving the ND 3 ؉ terminus that may contribute to the unusual thermostability of this molecule.
Journal of the American Chemical Society, 1998
High-level, all-electron, density functional calculations on a 104-atom model (B3LYP/6-311G** level) have been used, in conjunction with high-resolution X-ray structural data, to predict the remarkable paramagnetic contact shifts recently measured for 1 H, 2 H, 13 C, and 15 N nuclei in Clostridium pasteurianum rubredoxin. Three published X-ray structures for the Fe(III) rubredoxin from C. pasteurianum were employed to construct a 104-atom model for the iron center that included all atoms shown to have strong electronic interactions with the Fe. Each of these models served as a starting point for quantum mechanical calculations at level B3LYP/6-311G**, which, in turn, yielded calculated values for Fermi contact spin densities. The results indicate that the experimental hyperfine shifts are dominated by Fermi contact interactions: calculated Fermi contact spin densities were found to correlate linearly with isotropic hyperfine 1 H, 2 H, 13 C, and 15 N NMR chemical shifts determined for Fe(III) rubredoxin. At the current level of hyperfine peak assignments (all signals assigned to residue and atom types; some assigned to sequence specifically), comparisons were made between experimental shifts and those calculated from the structural model derived from each of the three X-ray structures. For Fe(III) rubredoxin, the R 2 values for the correlation between the calculated spin densities and experimental chemical shifts ranged, depending on the model, from 0.93 to 0.96 for 12 experimental 2 H signals and from 0.85 to 0.96 for 12 experimental 15 N signals. The correlation with experiment was improved by performing partial geometry optimizations at B3LYP/3-21G* on two of the 104-atom models. In particular, the R 2 correlation with experimental 15 N chemical shifts was improved from 0.85 to 0.94 upon optimizing the positions of the nitrogen bound protons reported for one of the X-ray structures. A small increase in the correlation with experiment was also found after optimizing the positions of the Rand -protons of the cysteines of another model. The consistent overall agreement between calculation and experiment supports the validity and usefulness of combining quantum chemical methods with NMR spectroscopy and X-ray crystallography for the testing and refinement of molecular structures. Significantly poorer correlations with experiment were obtained when hypothetical Fe(II) models, derived from two of the X-ray structures of Fe(III) rubredoxin, were used as the basis for the spin density calculations; this suggests that the protein undergoes subtle structural changes upon reduction. † Coordinates for the model structures, the input data for the Gaussian 94 calculations, and the calculated values for the Fermi contact spin densities have been deposited at BioMagResBank (URL: http://www.bmrb.wisc.edu) under the following accession numbers (in parentheses): oxidized rubredoxin models (4118); reduced rubredoxin models (4119). Except as noted above, all calculations were performed at B3LYP/6-311G**.
Pyrococcus furiosus is a hyperthermophilic Archaea. An uncharacterized protein of this Achaea, I6U7D0 (UniProt accession) containing 349 residues was selected for in silico analysis. Various bioinformatic tools were used to predict the structure and function of this protein. Sequence similarity was searched through UniProt and non-redundant database using BLASTp program of NCBI and homology was found with methyltransferases. Multiple sequence alignment was used to locate the conserved residues. The secondary and three dimensional structures were predicted. The validation of the three dimensional structure was obtained through PROCHECK, Verify3D and ERRAT program. CASTp server was used to predict the active site of the protein. Molecular docking with the ligand ACY (Acetic Acid) was performed using Molegro Virtual Docker to visualize the interactions between the ligand and amino acid residues in the protein. Finally, all the accumulated results suggested the biological function of the target protein to be a methyltransferase
Biochemical and Biophysical Research Communications, 2003
Rubredoxin (D.g. Rd), a small non-heme iron-sulfur protein shown to function as a redox coupling protein from the sulfate reducing bacteria Desulfovibrio gigas, has been crystallized using the hanging-drop vapor diffusion method and macroseeding method. Rubredoxin crystals diffract to an ultra-high resolution 0.68 A A using synchrotron radiation X-ray, and belong to the space group P2 1 with unit-cell parameters a ¼ 19:44 A A, b ¼ 41:24 A A, c ¼ 24:10 A A, and b ¼ 108:46°. The data set of single-wavelength anomalous dispersion signal of iron in the native crystal was also collected for ab initio structure re-determination. Preliminary analysis indicates that there is one monomer with a [Fe-4S] cluster in each asymmetric unit. The crystal structure at this ultra-high resolution will reveal the details of its biological function. The crystal character and data collection strategy for ultra-high resolution will also be discussed.