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1.
Karin E. van Straaten Claudio F. Gonzalez Ricardo B. Valladares Xiaohui Xu Alexei V. Savchenko David A. R. Sanders 《Protein science : a publication of the Protein Society》2009,18(10):2196-2202
The structure of the Atu1476 protein from Agrobacterium tumefaciens was determined at 2 Å resolution. The crystal structure and biochemical characterization of this enzyme support the conclusion that this protein is an S-formylglutathione hydrolase (AtuSFGH). The three-dimensional structure of AtuSFGH contains the α/β hydrolase fold topology and exists as a homo-dimer. Contacts between the two monomers in the dimer are formed both by hydrogen bonds and salt bridges. Biochemical characterization reveals that AtuSFGH hydrolyzes C—O bonds with high affinity toward short to medium chain esters, unlike the other known SFGHs which have greater affinity toward shorter chained esters. A potential role for Cys54 in regulation of enzyme activity through S-glutathionylation is also proposed. 相似文献
2.
Protein side chains make most of the specific contacts between proteins and other molecules, and their conformational properties have been studied for many years. These properties have been analyzed primarily in the form of rotamer libraries, which cluster the observed conformations into groups and provide frequencies and average dihedral angles for these groups. In recent years, these libraries have improved with higher resolution structures and using various criteria such as high thermal factors to eliminate side chains that may be misplaced within the crystallographic model coordinates. Many of these side chains have highly non-rotameric dihedral angles. The origin of side chains with high B-factors and/or with non-rotameric dihedral angles is of interest in the determination of protein structures and in assessing the prediction of side chain conformations. In this paper, using a statistical analysis of the electron density of a large set of proteins, it is shown that: (1) most non-rotameric side chains have low electron density compared to rotameric side chains; (2) up to 15% of chi1 non-rotameric side chains in PDB models can clearly be fit to density at a single rotameric conformation and in some cases multiple rotameric conformations; (3) a further 47% of non-rotameric side chains have highly dispersed electron density, indicating potentially interconverting rotameric conformations; (4) the entropy of these side chains is close to that of side chains annotated as having more than one chi(1) rotamer in the crystallographic model; (5) many rotameric side chains with high entropy clearly show multiple conformations that are not annotated in the crystallographic model. These results indicate that modeling of side chains alternating between rotamers in the electron density is important and needs further improvement, both in structure determination and in structure prediction. 相似文献
3.
Samantha Perez-Miller Qin Zou Milos V Novotny Thomas D Hurley 《Protein science : a publication of the Protein Society》2010,19(8):1469-1479
In mice, the major urinary proteins (MUP) play a key role in pheromonal communication by binding and transporting semiochemicals. MUP‐IV is the only isoform known to be expressed in the vomeronasal mucosa. In comparison with the MUP isoforms that are abundantly excreted in the urine, MUP‐IV is highly specific for the male mouse pheromone 2‐sec‐butyl‐4,5‐dihydrothiazole (SBT). To examine the structural basis of this ligand preference, we determined the X‐ray crystal structure of MUP‐IV bound to three mouse pheromones: SBT, 2,5‐dimethylpyrazine, and 2‐heptanone. We also obtained the structure of MUP‐IV with 2‐ethylhexanol bound in the cavity. These four structures show that relative to the major excreted MUP isoforms, three amino acid substitutions within the binding calyx impact ligand coordination. The F103 for A along with F54 for L result in a smaller cavity, potentially creating a more closely packed environment for the ligand. The E118 for G substitution introduces a charged group into a hydrophobic environment. The sidechain of E118 is observed to hydrogen bond to polar groups on all four ligands with nearly the same geometry as seen for the water‐mediated hydrogen bond network in the MUP‐I and MUP‐II crystal structures. These differences in cavity size and interactions between the protein and ligand are likely to contribute to the observed specificity of MUP‐IV. 相似文献
4.
SF3a is an evolutionarily conserved heterotrimeric complex essential for pre-mRNA splicing. It functions in spliceosome assembly within the mature U2 snRNP (small nuclear ribonucleoprotein particle), and its displacement from the spliceosome initiates the first step of the splicing reaction. We have identified a core domain of the yeast SF3a complex required for complex assembly and determined its crystal structure. The structure shows a bifurcated assembly of three subunits, Prp9, Prp11 and Prp21, with Prp9 interacting with Prp21 via a bidentate-binding mode, and Prp21 wrapping around Prp11. Structure-guided biochemical analysis also shows that Prp9 harbours a major binding site for stem-loop IIa of U2 snRNA. These findings provide mechanistic insights into the assembly of U2 snRNP. 相似文献
5.
Voronov-Goldman M Lamed R Noach I Borovok I Kwiat M Rosenheck S Shimon LJ Bayer EA Frolow F 《Proteins》2011,79(1):50-60
6.
The relative stability of protein structures determined by either X-ray crystallography or nuclear magnetic resonance (NMR) spectroscopy has been investigated by using molecular dynamics simulation techniques. Published structures of 34 proteins containing between 50 and 100 residues have been evaluated. The proteins selected represent a mixture of secondary structure types including all alpha, all beta, and alpha/beta. The proteins selected do not contain cysteine-cysteine bridges. In addition, any crystallographic waters, metal ions, cofactors, or bound ligands were removed before the systems were simulated. The stability of the structures was evaluated by simulating, under identical conditions, each of the proteins for at least 5 ns in explicit solvent. It is found that not only do NMR-derived structures have, on average, higher internal strain than structures determined by X-ray crystallography but that a significant proportion of the structures are unstable and rapidly diverge in simulations. 相似文献
7.
Yan Wang Jouko Virtanen Zhidong Xue John J. G. Tesmer Yang Zhang 《Acta Crystallographica. Section D, Structural Biology》2016,72(5):616-628
Molecular replacement (MR) often requires templates with high homology to solve the phase problem in X‐ray crystallography. I‐TASSER‐MR has been developed to test whether the success rate for structure determination of distant‐homology proteins could be improved by a combination of iterative fragmental structure‐assembly simulations with progressive sequence truncation designed to trim regions with high variation. The pipeline was tested on two independent protein sets consisting of 61 proteins from CASP8 and 100 high‐resolution proteins from the PDB. After excluding homologous templates, I‐TASSER generated full‐length models with an average TM‐score of 0.773, which is 12% higher than the best threading templates. Using these as search models, I‐TASSER‐MR found correct MR solutions for 95 of 161 targets as judged by having a TFZ of >8 or with the final structure closer to the native than the initial search models. The success rate was 16% higher than when using the best threading templates. I‐TASSER‐MR was also applied to 14 protein targets from structure genomics centers. Seven of these were successfully solved by I‐TASSER‐MR. These results confirm that advanced structure assembly and progressive structural editing can significantly improve the success rate of MR for targets with distant homology to proteins of known structure. 相似文献
8.
9.
Akifumi Higashiura Takeshi Kurakane Makoto Matsuda Mamoru Suzuki Koji Inaka Masaru Sato Tomoyuki Kobayashi Tetsuo Tanaka Hiroaki Tanaka Kazuko Fujiwara Atsushi Nakagawa 《Acta Crystallographica. Section D, Structural Biology》2010,66(6):698-708
Recent technical improvements in macromolecular X‐ray crystallography have significantly improved the resolution limit of protein structures. However, examples of high‐resolution structure determination are still limited. In this study, the X‐ray crystal structure of bovine H‐protein, a component of the glycine cleavage system, was determined at 0.88 Å resolution. This is the first ultrahigh‐resolution structure of an H‐protein. The data were collected using synchrotron radiation. Because of limitations of the hardware, especially the dynamic range of the CCD detector, three data sets (high‐, medium‐ and low‐resolution data sets) were measured in order to obtain a complete set of data. To improve the quality of the merged data, the reference data set was optimized for merging and the merged data were assessed by comparing merging statistics and R factors against the final model and the number of visualized H atoms. In addition, the advantages of merging three data sets were evaluated. The omission of low‐resolution reflections had an adverse effect on visualization of H atoms in hydrogen‐omit maps. Visualization of hydrogen electron density is a good indicator for assessing the quality of high‐resolution X‐ray diffraction data. 相似文献
10.
A systematic method for the analysis of the hydration structure of proteins is demonstrated on the case study of lysozyme. The method utilises multiple structural data of the same protein deposited in the protein data bank. Clusters of high water occupancy are localised and characterised in terms of their interaction with protein. It is shown that they constitute a network of interconnected hydrogen bonds anchored to the protein molecule. The high occupancy of the clusters does not directly correlate with water–protein interaction energy as was originally hypothesised. The highly occupied clusters rather correspond to the nodes of the hydration network that have the maximum number of hydrogen bonds including both the protein atoms and the surrounding water clusters. Copyright © 2013 John Wiley & Sons, Ltd. 相似文献
11.
John H. Beale 《Acta Crystallographica. Section D, Structural Biology》2020,76(5):400-405
The number of new X‐ray crystallography‐based submissions to the Protein Data Bank appears to be at the beginning of a decline, perhaps signalling an end to the era of the dominance of X‐ray crystallography within structural biology. This letter, from the viewpoint of a young structural biologist, applies the Copernican method to the life expectancy of crystallography and asks whether the technique is still the mainstay of structural biology. A study of the rate of Protein Data Bank depositions allows a more nuanced analysis of the fortunes of macromolecular X‐ray crystallography and shows that cryo‐electron microscopy might now be outcompeting crystallography for new labour and talent, perhaps heralding a change in the landscape of the field. 相似文献
12.
The three-dimensional structure of a mutant human lysozyme, W64CC65A, in which a non-native disulfide bond Cys64--Cys81 is substituted for the Cys65--Cys81 of the wild type protein by replacing Trp64 and Cys65 with Cys and Ala, respectively, was determined by X-ray crystallography and refined to an R-value of 0.181, using 33,187 reflections at 1.87-A resolution. The refined model of the W64CC65A protein consisted of four molecules, which were related by two noncrystallographic twofold axes and a translation vector. Although no specific structural differences could be observed among these four molecules, the overall B-factors of each molecule were quite different. The overall structure of W64CC65A, especially in the alpha-helical domain, was found to be quite similar to that of the wild type protein. Moreover, the side-chain conformation of the newly formed Cys64--Cys81 bond was quite similar to that of the Cys65--Cys81 bond of the wild-type protein. However, in the beta-sheet domain, the main-chain atoms of the loop region from positions 66-75 could not be determined, and significant structural changes due to the formation of the non-native disulfide bond could be observed. From these results, it is clear that the loop region of the mutant protein does not fold with the specific folding as observed in the wild-type protein. 相似文献
13.
The 2.1-A resolution crystal structure of the MATalpha2 homeodomain bound to DNA reveals the unexpected presence of two nonspecifically bound alpha2 homeodomains, in addition to the two alpha2 homeodomains bound to canonical alpha2 binding sites. One of the extra homeodomains makes few base-specific contacts, while the other extra homeodomain binds to DNA in a previously unobserved manner. This unusually bound homeodomain is rotated on the DNA, making possible major groove contacts by side-chains that normally do not contact the DNA. This alternate docking may represent one way in which homeodomains sample nonspecific DNA sequences. 相似文献
14.
Sarah Triest Alexandre Wohlknig Els Pardon Jan Steyaert 《Acta Crystallographica. Section F, Structural Biology Communications》2014,70(11):1504-1507
GPCR–G‐protein complexes are one of the most important components of cell‐signalling cascades. Extracellular signals are sensed by membrane‐associated G‐protein‐coupled receptors (GPCRs) and transduced via G proteins towards intracellular effector molecules. Structural studies of these transient complexes are crucial to understand the molecular details of these interactions. Although a nucleotide‐free GPCR–G‐protein complex is stable, it is not an ideal sample for crystallization owing to the intrinsic mobility of the Gαs α‐helical domain (AHD). To stabilize GPCR–G‐protein complexes in a nucleotide‐free form, nanobodies were selected that target the flexible GαsAHD. One of these nanobodies, CA9177, was co‐crystallized with the GαsAHD. Initial crystals were obtained using the sitting‐drop method in a sparse‐matrix screen and further optimized. The crystals diffracted to 1.59 Å resolution and belonged to the monoclinic space group P21, with unit‐cell parameters a = 44.07, b = 52.55, c = 52.66 Å, α = 90.00, β = 107.89, γ = 90.00°. The structure of this specific nanobody reveals its binding epitope on GαsAHD and will help to determine whether this nanobody could be used as crystallization chaperone for GPCR–G‐protein complexes. 相似文献
15.
We determined the three-dimensional (3D) crystal structure of protein TM841, a protein product from a hypothetical open-reading frame in the genome of the hyperthermophile bacterium Thermotoga maritima, to 2.0 A resolution. The protein belongs to a large protein family, DegV or COG1307 of unknown function. The 35 kDa protein consists of two separate domains, with low-level structural resemblance to domains from other proteins with known 3D structures. These structural homologies, however, provided no clues for the function of TM841. But the electron density maps revealed clear density for a bound fatty-acid molecule in a pocket between the two protein domains. The structure indicates that TM841 has the molecular function of fatty-acid binding and may play a role in the cellular functions of fatty acid transport or metabolism. 相似文献
16.
Corazza A Rosano C Pagano K Alverdi V Esposito G Capanni C Bemporad F Plakoutsi G Stefani M Chiti F Zuccotti S Bolognesi M Viglino P 《Proteins》2006,62(1):64-79
The structure of AcP from the hyperthermophilic archaeon Sulfolobus solfataricus has been determined by (1)H-NMR spectroscopy and X-ray crystallography. Solution and crystal structures (1.27 A resolution, R-factor 13.7%) were obtained on the full-length protein and on an N-truncated form lacking the first 12 residues, respectively. The overall Sso AcP fold, starting at residue 13, displays the same betaalphabetabetaalphabeta topology previously described for other members of the AcP family from mesophilic sources. The unstructured N-terminal tail may be crucial for the unusual aggregation mechanism of Sso AcP previously reported. Sso AcP catalytic activity is reduced at room temperature but rises at its working temperature to values comparable to those displayed by its mesophilic counterparts at 25-37 degrees C. Such a reduced activity can result from protein rigidity and from the active site stiffening due the presence of a salt bridge between the C-terminal carboxylate and the active site arginine. Sso AcP is characterized by a melting temperature, Tm, of 100.8 degrees C and an unfolding free energy, DeltaG(U-F)H2O, at 28 degrees C and 81 degrees C of 48.7 and 20.6 kJ mol(-1), respectively. The kinetic and structural data indicate that mesophilic and hyperthermophilic AcP's display similar enzymatic activities and conformational stabilities at their working conditions. Structural analysis of the factor responsible for Sso AcP thermostability with respect to mesophilic AcP's revealed the importance of a ion pair network stabilizing particularly the beta-sheet and the loop connecting the fourth and fifth strands, together with increased density packing, loop shortening and a higher alpha-helical propensity. 相似文献
17.
Soraya de Chadarevian 《Protein science : a publication of the Protein Society》2018,27(6):1136-1143
The essay reviews John Kendrew's pioneering work on the structure of myoglobin for which he shared the Nobel Prize for Chemistry in 1962. It reconstructs the status of protein X‐ray crystallography at the time Kendrew entered the field in 1945, after distinctive service in operational research during the war. It reflects on the choice of sperm whale myoglobin as research material. In particular, it highlights Kendrew's early use of digital electronic computers for crystallographic computations and the marshaling of other tools and approaches that made it possible to solve the structure at increasing resolution. The essay further discusses the role of models in structure resolution and their broader reception. It ends by briefly reviewing Kendrew's other contributions in the formation and institutionalization of molecular biology. 相似文献
18.
Exploration of disorder in protein structures by X-ray restrained molecular dynamics 总被引：2，自引：0，他引：2
Conformational disorder in crystal structures of ribonuclease-A and crambin is studied by including two independent structures in least-squares optimizations against X-ray data. The optimizations are carried out by X-ray restrained molecular dynamics (simulated annealing refinement) and by conventional least-squares optimization. Starting from two identical structures, the optimizations against X-ray data lead to significant deviations between the two, with rms backbone displacements of 0.45 A for refinement of ribonuclease at 1.53 A resolution, and 0.31 A for crambin at 0.945 A. More than 15 independent X-ray restrained molecular dynamics runs have been carried out for ribonuclease, and the displacements between the resulting structures are highly reproducible for most atoms. These include residues with two or more conformations with significant dihedral angle differences and alternative hydrogen bonding, as well as groups of residues that undergo displacements that are suggestive of rigid-body librations. The crystallographic R-values obtained are approximately 13%, as compared to 15.3% for a comparable refinement with a single structure. Least-squares optimization without an intervening restrained molecular dynamics stage is sufficient to reproduce most of the observed displacements. Similar results are obtained for crambin, where the higher resolution of the X-ray data allows for refinement of unconstrained individual anisotropic temperature factors. These are shown to be correlated with the displacements in the two-structure refinements. 相似文献
19.
20.
Jin Zhang Katsuhide Mizuno Yuki Murata Hideaki Koide Midori Murakami Kunio Ihara Tsutomu Kouyama 《Proteins》2013,81(9):1585-1592
Deltarhodopsin, a new member of the microbial rhodopsin family, functions as a light‐driven proton pump. Here, we report the three‐dimensional structure of deltarhodopsin (dR3) from Haloterrigena thermotolerans at 2.7 Å resolution. A crystal belonging to space group R32 (a, b = 111.71 Å, c = 198.25 Å) was obtained by the membrane fusion method. In this crystal, dR3 forms a trimeric structure as observed for bacteriorhodopsin (bR). Structural comparison of dR with bR showed that the inner part (the proton release and uptake pathways) is highly conserved. Meanwhile, residues in the protein–protein contact region are largely altered so that the diameter of the trimeric structure at the cytoplasmic side is noticeably larger in dR3. Unlike bR, dR3 possesses a helical segment at the C‐terminal region that fills the space between the AB and EF loops. A significant difference is also seen in the FG loop, which is one residue longer in dR3. Another peculiar property of dR3 is a highly crowded distribution of positively charged residues on the cytoplasmic surface, which may be relevant to a specific interaction with some cytoplasmic component.Proteins 2013; © 2013 Wiley Periodicals, Inc. 相似文献