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Anomalous Diffraction (anomalous + diffraction)
Kinds of Anomalous Diffraction Selected AbstractsStructures of Four Crystal Forms of DecaplaninHELVETICA CHIMICA ACTA, Issue 5 2003Christopher Lehmann The glycopeptide antibiotic decaplanin (1; formerly known as MM 47761 and M86-1410) crystallizes in two P21 and two P6122 crystal forms, each with four monomers in the asymmetric unit, with solvent contents varying from 48 to 69%. Although with ca. 600 unique atoms, the structures are larger than typical small molecules, one was solved by direct methods. The other three were solved by typical macromolecular methods: single-wavelength anomalous diffraction (SAD) of the Cl-atoms present naturally in the structure, multiple-wavelength anomalous diffraction (MAD) at the Br absorption edge for a crystal soaked in NaBr solution, and molecular replacement. There is evidence of appreciable radiation damage with loss of 20,30% of the covalent and ionic halogens affecting the synchrotron datasets that may even have unintentionally facilitated the MAD structure solution. The structures contain the dimer units typical of antibiotics related to vancomycin, but, in addition, there are a variety of further intermolecular interactions responsible for the polymorphy leading to intertwined 61 -helices in two of the crystal forms. Except for the sugars and some sidechains, the conformations of the 16 independent monomers are very similar. [source] The revenge of the Patterson methods.JOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 2 2007The Patterson techniques, recently developed by the same authors for the ab initio crystal structure solution of proteins, have been applied to single and multiple anomalous diffraction (SAD and MAD) data to find the substructure of the anomalous scatterers. An automatic procedure has been applied to a large set of test structures, some of which were originally solved with remarkable difficulty. In all cases, the procedure automatically leads to interpretable electron density maps. Patterson techniques have been compared with direct methods; the former seem to be more efficient than the latter, so confirming the results obtained for ab initio phasing, and disproving the common belief that they could only be applied to determine large equal-atom substructures with difficulty. [source] A parallel program using SHELXD for quick heavy-atom partial structural solution on high-performance computersJOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 2 2007Zheng-Qing Fu A parallel algorithm has been designed for SHELXD to solve the heavy-atom partial structures of protein crystals quickly. Based on this algorithm, a program has been developed to run on high-performance multiple-CPU Linux PCs, workstations or clusters. Tests on the 32-CPU Linux cluster at SER-CAT, APS, Argonne National Laboratory, show that the parallelization dramatically speeds up the process by a factor of roughly the number of CPUs applied, leading to reliable and instant heavy-atom sites solution, which provides the practical opportunity to employ heavy-atom search as an alternative tool for anomalous scattering data quality evaluation during single/multiple-wavelength anomalous diffraction (SAD/MAD) data collection at synchrotron beamlines. [source] Evaluation of SnB for the location of anomalous scattering atoms in SAD/MAD phasingJOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 6 2003Jun Wang Sixteen existing single-wavelength/multi-wavelength anomalous diffraction (SAD/MAD) data sets with a broad range of crystallographic properties have been used to investigate the various parameters in SnB with the goal of finding the optimum values for locating the positions of anomalous scatterers. The results of the analysis indicate some changes in default parameters that may be useful for non-routine and difficult cases. [source] Phasing possibilities using different wavelengths with a xenon derivativeJOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 2 2002Santosh Panjikar Xenon derivatives are generally expected to be isomorphous with the native; however, the K - and L -absorption edges are not easily accessible on most synchrotron beamlines, which might limit their usefulness in phase determination. Various phasing procedures for xenon-derivatized porcine pancreatic elastase have been investigated using data sets measured at three generally accessible wavelengths. The importance of highly redundant data in measuring precise anomalous differences is highlighted and it is shown that, after such measurements, a single isomorphous replacement anomalous scattering (SIRAS) procedure yields a better phase set than those generated by single anomalous scattering (SAS) or multiwavelength anomalous diffraction (MAD) procedures. [source] The X-ray crystal structure of PA1607 from Pseudomonas aureginosa at 1.9 Å resolution,a putative transcription factorPROTEIN SCIENCE, Issue 3 2007Edyta A.L. Sieminska Abstract The structure of the PA1607 protein from Pseudomonas aureginosa was determined at 1.85 Å resolution using the Se-Met multiwavelength anomalous diffraction (MAD) technique. PA1607 forms a dimer and adopts a winged-helix motif similar to the MarR family of transcription regulators, though it has an unusual dimerization profile. The DNA-binding regions and a putative metal-binding site are not conserved in PA1607. [source] Enhancing MAD FA data for substructure determinationACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2010Hongliang Xu Heavy-atom substructure determination is a critical step in phasing an unknown macromolecular structure. Dual-space (Shake-and-Bake) recycling is a very effective procedure for locating the substructure (heavy) atoms using FA data estimated from multiple-wavelength anomalous diffraction. However, the estimated FA are susceptible to the accumulation of errors in the individual intensity measurements at several wavelengths and from inaccurate estimation of the anomalous atomic scattering corrections f, and f,,. In this paper, a new statistical and computational procedure which merges multiple FA estimates into an averaged data set is used to further improve the quality of the estimated anomalous amplitudes. The results of 18 Se-atom substructure determinations provide convincing evidence in favor of using such a procedure to locate anomalous scatterers. [source] A dipicolinate lanthanide complex for solving protein structures using anomalous diffractionACTA CRYSTALLOGRAPHICA SECTION D, Issue 7 2010Guillaume Pompidor Tris-dipicolinate lanthanide complexes were used to prepare derivative crystals of six proteins: hen egg-white lysozyme, turkey egg-white lysozyme, thaumatin from Thaumatococcus daniellii, urate oxidase from Aspergillus flavus, porcine pancreatic elastase and xylanase from Trichoderma reesei. Diffraction data were collected using either synchrotron radiation or X-rays from a laboratory source. In all cases, the complex turned out to be bound to the protein and the phases determined using the anomalous scattering of the lanthanide led to high-quality electron-density maps. The binding mode of the complex was characterized from the refined structures. The lanthanide tris-dipicolinate was found to bind through interactions between carboxylate groups of the dipicolinate ligands and hydrogen-bond donor groups of the protein. In each binding site, one enantiomeric form of the complex is selected from the racemic solution according to the specific site topology. For hen egg-white lysozyme and xylanase, derivative crystals obtained by cocrystallization belonged to a new monoclinic C2 crystal form that diffracted to high resolution. [source] Structural characterization of tartrate dehydrogenase: a versatile enzyme catalyzing multiple reactionsACTA CRYSTALLOGRAPHICA SECTION D, Issue 6 2010Radhika Malik The first structure of an NAD-dependent tartrate dehydrogenase (TDH) has been solved to 2,Å resolution by single anomalous diffraction (SAD) phasing as a complex with the intermediate analog oxalate, Mg2+ and NADH. This TDH structure from Pseudomonas putida has a similar overall fold and domain organization to other structurally characterized members of the hydroxy-acid dehydrogenase family. However, there are considerable differences between TDH and these functionally related enzymes in the regions connecting the core secondary structure and in the relative positioning of important loops and helices. The active site in these complexes is highly ordered, allowing the identification of the substrate-binding and cofactor-binding groups and the ligands to the metal ions. Residues from the adjacent subunit are involved in both the substrate and divalent metal ion binding sites, establishing a dimer as the functional unit and providing structural support for an alternating-site reaction mechanism. The divalent metal ion plays a prominent role in substrate binding and orientation, together with several active-site arginines. Functional groups from both subunits form the cofactor-binding site and the ammonium ion aids in the orientation of the nicotinamide ring of the cofactor. A lysyl amino group (Lys192) is the base responsible for the water-mediated proton abstraction from the C2 hydroxyl group of the substrate that begins the catalytic reaction, followed by hydride transfer to NAD. A tyrosyl hydroxyl group (Tyr141) functions as a general acid to protonate the enolate intermediate. Each substrate undergoes the initial hydride transfer, but differences in substrate orientation are proposed to account for the different reactions catalyzed by TDH. [source] Getting the best out of long-wavelength X-rays: de novo chlorine/sulfur SAD phasing of a structural protein from ATVACTA CRYSTALLOGRAPHICA SECTION D, Issue 3 2010Adeline Goulet The structure of a 14,kDa structural protein from Acidianus two-tailed virus (ATV) was solved by single-wavelength anomalous diffraction (SAD) phasing using X-ray data collected at 2.0,Å wavelength. Although the anomalous signal from methionine sulfurs was expected to suffice to solve the structure, one chloride ion turned out to be essential to achieve phasing. The minimal data requirements and the relative contributions of the Cl and S atoms to phasing are discussed. This work supports the feasibility of a systematic approach for the solution of protein crystal structures by SAD based on intrinsic protein light atoms along with associated chloride ions from the solvent. In such cases, data collection at long wavelengths may be a time-efficient alternative to selenomethionine substitution and heavy-atom derivatization. [source] Novel structural features in the GMC family of oxidoreductases revealed by the crystal structure of fungal aryl-alcohol oxidaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2009Israel S. Fernández Lignin biodegradation, a key step in carbon recycling in land ecosystems, is carried out by white-rot fungi through an H2O2 -dependent process defined as enzymatic combustion. Pleurotus eryngii is a selective lignin-degrading fungus that produces H2O2 during redox cycling of p -anisylic compounds involving the secreted flavoenzyme aryl-alcohol oxidase (AAO). Here, the 2.4,Å resolution X-ray crystal structure of this oxidoreductase, which catalyzes dehydrogenation reactions on various primary polyunsaturated alcohols, yielding the corresponding aldehydes, is reported. The AAO crystal structure was solved by single-wavelength anomalous diffraction of a selenomethionine derivative obtained by Escherichia coli expression and in vitro folding. This monomeric enzyme is composed of two domains, the overall folding of which places it into the GMC (glucose,methanol,choline oxidase) oxidoreductase family, and a noncovalently bound FAD cofactor. However, two additional structural elements exist in the surroundings of its active site that modulate the access of substrates; these are absent in the structure of the model GMC oxidoreductase glucose oxidase. The folding of these novel elements gives rise to a funnel-like hydrophobic channel that connects the solvent region to the buried active-site cavity of AAO. This putative active-site cavity is located in front of the re side of the FAD isoalloxazine ring and near two histidines (His502 and His546) that could contribute to alcohol activation as catalytic bases. Moreover, three aromatic side chains from two phenylalanines (Phe397 and Phe502) and one tyrosine (Tyr92) at the inner region of the channel form an aromatic gate that may regulate the access of the enzyme substrates to the active site as well as contribute to the recognition of the alcohols that can effectively be oxidized by AAO. [source] Direct-method SAD phasing of proteins enhanced by the use of intrinsic bimodal phase distributions in the subsequent phase-improvement processACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2009Li-Jie Wu A modified SAD (single-wavelength anomalous diffraction) phasing algorithm has been introduced in the latest version of the program OASIS. In addition to direct-method phases and figures of merit, Hendrickson,Lattman coefficients that correspond to the original unresolved bimodal phase distributions are also output and used in subsequent phase-improvement procedures in combination with the improved phases. This provides the possibility of rebreaking the SAD phase ambiguity using the ever-improving phases resulting from the phase-improvement process. Tests using experimental SAD data from six known proteins showed that in all cases the new treatment produced significant improved results. [source] On the combination of molecular replacement and single-wavelength anomalous diffraction phasing for automated structure determinationACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2009Santosh Panjikar A combination of molecular replacement and single-wavelength anomalous diffraction phasing has been incorporated into the automated structure-determination platform Auto-Rickshaw. The complete MRSAD procedure includes molecular replacement, model refinement, experimental phasing, phase improvement and automated model building. The improvement over the standard SAD or MR approaches is illustrated by ten test cases taken from the JCSG diffraction data-set database. Poor MR or SAD phases with phase errors larger than 70° can be improved using the described procedure and a large fraction of the model can be determined in a purely automatic manner from X-ray data extending to better than 2.6,Å resolution. [source] Structure of 3-deoxy- manno -octulosonate cytidylyltransferase from Haemophilus influenzae complexed with the substrate 3-deoxy- manno -octulosonate in the ,-configurationACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2008Hye-Jin Yoon The enzyme 3-deoxy- manno -octulosonate cytidylyltransferase (CMP-KDO synthetase; CKS) catalyzes the activation of 3-deoxy- d - manno -octulosonate (or 2-keto-3-deoxy- manno -octonic acid; KDO) by forming CMP-KDO. CKS is unique to Gram-negative bacteria and is an attractive target for the development of antibacterial agents. The crystal structure of CKS from Haemophilus influenzae in complex with the substrate KDO has been determined at 2.30,Å resolution by combining single-wavelength anomalous diffraction and molecular-replacement methods. The two monomers in the asymmetric unit differ in the conformation of their C-terminal ,-helix (Ala230,Asn254). The KDO bound to the active site exists as the ,-pyranose form in the 5C2 chair conformation. The structure of CKS from H. influenzae in complex with KDO will be useful in structure-based inhibitor design. [source] Making the most of two crystals: structural analysis of a conserved hypothetical protein using native gel screening and SAD phasingACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2003J. Shaun Lott The protein PAE2307 is a member of a protein family of unknown function which is conserved among a number of bacterial and archaeal species. The protein was overexpressed in Escherichia coli, purified and crystallized in two crystal forms. The prevalent form was twinned, but the other diffracted to 1.45,Å resolution. The non-twinned crystals proved difficult to reproduce, so screening of potential heavy-atom derivatives by native polyacrylamide gel electrophoresis was used to establish suitable derivatization conditions. This process enabled the production of a K2Pt(NO2)4 derivative that was used to collect a single-wavelength anomalous diffraction (SAD) data set from the only available crystal. Phase information of high quality was obtained, enabling the calculation of an interpretable electron-density map. [source] Twinned crystals and anomalous phasingACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2003Zbigniew Dauter Merohedral or pseudomerohedral twinning of crystals cannot be identified from inspection of the diffraction patterns. Several methods for the identification of twinning and the estimation of the twin fraction are suitable for macromolecular crystals and all are based on the statistical properties of the measured diffraction intensities. If the crystal twin fraction is estimated and is not too close to 0.5, the diffraction data can be detwinned; that is, related to the individual crystal specimen. However, the detwinning procedure invariably introduces additional inaccuracies to the estimated intensities, which substantially increase when the twin fraction approaches 0.5. In some cases, a crystal structure can be solved with the original twinned data by standard techniques such as molecular replacement, multiple isomorphous replacement or multiwavelength anomalous diffraction. Test calculations on data collected from a twinned crystal of gpD, the bacteriophage , capsid protein, show that the single-wavelength anomalous diffraction (SAD) method can be used to solve its structure even if the data set corresponds to a perfectly twinned crystal with a twin fraction of 0.5. [source] High-phasing-power lanthanide derivatives: taking advantage of ytterbium and lutetium for optimized anomalous diffraction experiments using synchrotron radiationACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2003É. Girard Ytterbium and lutetium are well suited for optimized anomalous diffraction experiments using synchrotron radiation. Therefore, two lanthanide complexes Yb-HPDO3A and Lu-HPDO3A have been produced that are similar to the Gd-HPDO3A complex already known to give good derivative crystals. Derivative crystals of hen egg-white lysozyme were obtained by co-crystallization using 100,mM solutions of each lanthanide complex. De novo phasing has been carried out using single-wavelength anomalous diffraction on data sets collected on each derivative crystal at the LIII absorption edge of the corresponding lanthanide ( = 28,e,). A third data set was collected on a Lu-HPDO3A derivative crystal at the Se,K absorption edge with = 10,e,. The structures were refined and compared with the known structure of the Gd-HPDO3A lysozyme derivative. The quality of the experimental electron-density maps allows easy model building. With LIII absorption edges at shorter wavelengths than the gadolinium absorption edge, lutetium and ytterbium, when chelated by a ligand such as HPDO3A, form lanthanide complexes that are especially interesting for synchrotron-radiation experiments in structural biology. [source] Crystallization and rhenium MAD phasing of the acyl-homoserinelactone synthase EsaIACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2001William T. Watson Acyl-homoserine- l -lactones (AHLs) are diffusible chemical signals that are required for virulence of many Gram-negative bacteria. AHLs are produced by AHL synthases from two substrates, S -adenosyl- l -methionine and acyl-acyl carrier protein. The AHL synthase EsaI, which is homologous to the AHL synthases from other pathogenic bacterial species, has been crystallized in the primitive tetragonal space group P43, with unit-cell parameters a = b = 66.40, c = 47.33,Å. The structure was solved by multiple-wavelength anomalous diffraction with a novel use of the rhenium anomalous signal. The rhenium-containing structure has been refined to a resolution of 2.5,Å and the perrhenate ion binding sites and liganding residues have been identified. [source] Crystallization and preliminary X-ray diffraction analysis of a rat biliverdin reductaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2000Danyu Sun Biliverdin reductase (BVR) catalyzes the final step of haem degradation and converts biliverdin to bilirubin using NAD(P)H as an electron donor. This paper deals with the first crystallization and preliminary crystallographic study of recombinant rat BVR expressed in Escherichia coli. Crystals of BVR were obtained by the sitting-drop vapour-diffusion method. Using synchrotron radiation at station BL44B2 of SPring-8, Japan, BVR diffraction data were collected to 1.6,Å resolution. Crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 58.89, b = 70.41, c = 87.76,Å. The complete determination of the crystallographic structure is currently in progress using MAD (multiwavelength anomalous diffraction) data from an Ir-derivative crystal. [source] X-ray analysis of bilirubin oxidase from Myrothecium verrucaria at 2.3,Å resolution using a twinned crystalACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2010Kimihiko Mizutani Bilirubin oxidase (BOD), a multicopper oxidase found in Myrothecium verrucaria, catalyzes the oxidation of bilirubin to biliverdin. Oxygen is the electron acceptor and is reduced to water. BOD is used for diagnostic analysis of bilirubin in serum and has attracted considerable attention as an enzymatic catalyst for the cathode of biofuel cells that work under neutral conditions. Here, the crystal structure of BOD is reported for the first time. Blue bipyramid-shaped crystals of BOD obtained in 2-methyl-2,4-pentanediol (MPD) and ammonium sulfate solution were merohedrally twinned in space group P63. Structure determination was achieved by the single anomalous diffraction (SAD) method using the anomalous diffraction of Cu atoms and synchrotron radiation and twin refinement was performed in the resolution range 33,2.3,Å. The overall organization of BOD is almost the same as that of other multicopper oxidases: the protein is folded into three domains and a total of four copper-binding sites are found in domains 1 and 3. Although the four copper-binding sites were almost identical to those of other multicopper oxidases, the hydrophilic Asn residue (at the same position as a hydrophobic residue such as Leu in other multicopper oxidases) very close to the type I copper might contribute to the characteristically high redox potential of BOD. [source] Crystallization and preliminary X-ray studies of the N-domain of the Wilson disease associated proteinACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 6 2009Lili Liu Wilson disease associated protein (ATP7B) is essential for copper transport in human cells. Mutations that affect ATP7B function result in Wilson's disease, a chronic copper toxicosis. Disease-causing mutations within the N-domain of ATP7B (WND) are known to disrupt ATP binding, but a high-resolution X-ray structure of the ATP-binding site has not been reported. The N-domain was modified to delete the disordered loop comprising residues His1115,Asp1138 (WND,1115,1138). Unlike the wild-type N-domain, WND,1115,1138 formed good-quality crystals. Synchrotron diffraction data have been collected from WND,1115,1138 at the Canadian Light Source. A native WND,1115,1138 crystal diffracted to 1.7,Å resolution and belonged to space group P42212, with unit-cell parameters a = 39.2, b = 39.2, c = 168.9,Å. MAD data were collected to 2.7,Å resolution from a SeMet-derivative crystal with unit-cell parameters a = 38.4, b = 38.4, c = 166.7,Å. The WND,1115,1138 structure is likely to be solved by phasing from multiwavelength anomalous diffraction (MAD) experiments. [source] Structure of Lmaj006129AAA, a hypothetical protein from Leishmania majorACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2006Tracy Arakaki The gene product of structural genomics target Lmaj006129 from Leishmania major codes for a 164-residue protein of unknown function. When SeMet expression of the full-length gene product failed, several truncation variants were created with the aid of Ginzu, a domain-prediction method. 11 truncations were selected for expression, purification and crystallization based upon secondary-structure elements and disorder. The structure of one of these variants, Lmaj006129AAH, was solved by multiple-wavelength anomalous diffraction (MAD) using ELVES, an automatic protein crystal structure-determination system. This model was then successfully used as a molecular-replacement probe for the parent full-length target, Lmaj006129AAA. The final structure of Lmaj006129AAA was refined to an R value of 0.185 (Rfree = 0.229) at 1.60,Å resolution. Structure and sequence comparisons based on Lmaj006129AAA suggest that proteins belonging to Pfam sequence families PF04543 and PF01878 may share a common ligand-binding motif. [source] The structure at 2.5,Å resolution of human basophilic leukemia-expressed protein BLES03ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2005Eduard Bitto The crystal structure of the human basophilic leukemia-expressed protein (BLES03, p5326, Hs.433573) was determined by single-wavelength anomalous diffraction and refined to an R factor of 18.8% (Rfree = 24.5%) at 2.5,Å resolution. BLES03 shows no detectable sequence similarity to any functionally characterized proteins using state-of-the-art sequence-comparison tools. The structure of BLES03 adopts a fold similar to that of eukaryotic transcription initiation factor 4E (eIF4E), a protein involved in the recognition of the cap structure of eukaryotic mRNA. In addition to fold similarity, the electrostatic surface potentials of BLES03 and eIF4E show a clear conservation of basic and acidic patches. In the crystal lattice, the acidic amino-terminal helices of BLES03 monomers are bound within the basic cavity of symmetry-related monomers in a manner analogous to the binding of mRNA by eIF4E. Interestingly, the gene locus encoding BLES03 is located between genes encoding the proteins DRAP1 and FOSL1, both of which are involved in transcription initiation. It is hypothesized that BLES03 itself may be involved in a biochemical process that requires recognition of nucleic acids. [source] | ||||||||||