scholarly journals ANODE: anomalous and heavy-atom density calculation

2011 ◽  
Vol 44 (6) ◽  
pp. 1285-1287 ◽  
Author(s):  
Andrea Thorn ◽  
George M. Sheldrick
Keyword(s):  
Phase Shift ◽  
Heavy Atom ◽  
Data Bank ◽  
Protein Data Bank File ◽  
Anomalous Scattering ◽  
Atom Density ◽  
Usual Procedure ◽  
Isomorphous Replacement ◽  
Multiple Wavelength ◽  
Potential Applications

The new programANODEestimates anomalous or heavy-atom density by reversing the usual procedure for experimental phase determination by methods such as single- and multiple-wavelength anomalous diffraction and single isomorphous replacement anomalous scattering. Instead of adding a phase shift to the heavy-atom phases to obtain a starting value for the native protein phase, this phase shift is subtracted from the native phase to obtain the heavy-atom substructure phase. The required native phase is calculated from the information in a Protein Data Bank file of the structure. The resulting density enables even very weak anomalous scatterers such as sulfur to be located. Potential applications include the identification of unknown atoms and the validation of molecular replacement solutions.

scholarly journals HAD, a Data Bank of Heavy-Atom Binding Sites in Protein Crystals: a Resource for Use in Multiple Isomorphous Replacement and Anomalous Scattering

1998 ◽  
Vol 54 (6) ◽  
pp. 1199-1206 ◽  
Author(s):  
Suhail A. Islam ◽  
David Carvin ◽  
Michael J. E. Sternberg ◽  
Tom L. Blundell
Keyword(s):  
Binding Sites ◽  
Heavy Atom ◽  
Data Bank ◽  
Protein Crystals ◽  
Anomalous Scattering ◽  
Isomorphous Replacement ◽  
Crystallization Conditions ◽  
Derivatives Of ◽  
Coordinate Data

Information on the preparation and characterization of heavy-atom derivatives of protein crystals has been collected, either from the literature or directly from protein crystallographers, and assembled in the form of a heavy-atom data bank (HAD). The data bank contains coordinate data for the heavy-atom positions in a form that is compatible with the crystallographic data in the Brookhaven Protein Data Bank, together with a wealth of information on the crystallization conditions, the nature of the heavy-atom reagent and references to relevant publications. Some statistical information derived from the data bank, such as the most popular heavy-atom derivatives, is also included. The information can be directly accessed and should be useful to protein crystallographers seeking to improve their success in preparing heavy-atom derivatives for the methods of isomorphous replacement and anomalous dispersion. The World Wide Web address of HAD is http://www.icnet.uk/bmm/had.

Freeze-Trapping Isomorphous Xenon Derivatives of Protein Crystals

1997 ◽  
Vol 30 (4) ◽  
pp. 476-486 ◽  
Author(s):  
O. Sauer ◽  
A. Schmidt ◽  
C. Kratky
Keyword(s):  
Heavy Atom ◽  
Sperm Whale ◽  
Protein Crystal ◽  
Protein Crystals ◽  
Anomalous Scattering ◽  
Isomorphous Replacement ◽  
Potential Applications ◽  
Xenon Pressure ◽  
Gas Consumption ◽  
Derivatives Of

A simple and efficient method to prepare isomorphous derivatives of protein crystals with xenon as a heavy atom is described. The method consists of exposing a crystal to xenon gas of pressures above 5 atm (~ 0.5 MPa) for several minutes and subsequently shock-freezing the crystal to immobilize the xenon dissolved in the mother liquor and bound to the protein. Diffraction data can the be collected with the established techniques of protein cryocrystallography. Two types of high-pressure device are described to expose a protein crystal to the required xenon pressure, permitting rapid freeze-quenching after xenon exposure. One of these devices can be used for gas pressures up to and exceeding 5.0 MPa, with a gas consumption of a few millilitres of uncompressed gas. The technique has been tested with monoclinic crystals of sperm-whale metmyoglobin, which has four xenon binding sites. The results of these experiments are described and discussed. Potential applications of this technique include-besides the classical multiwavelength anomalous diffraction (MAD) or single isomorphous replacement with anomalous scattering (SIRAS) experiments-the derivation of low-angle phase information by modifying the electron density of the solvent regions within the crystal.

IL MILIONE: a suite of computer programs for crystal structure solution of proteins

2007 ◽  
Vol 40 (3) ◽  
pp. 609-613 ◽  
Author(s):  
Maria C. Burla ◽  
Rocco Caliandro ◽  
Mercedes Camalli ◽  
Benedetta Carrozzini ◽  
Giovanni L. Cascarano ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Electron Density ◽  
Heavy Atom ◽  
Direct Methods ◽  
Computer Programs ◽  
Atomic Resolution ◽  
Protein Crystal ◽  
Anomalous Scattering ◽  
Isomorphous Replacement ◽  
Phase Extension

IL MILIONEis a suite of computer programs devoted to protein crystal structure determination by X-ray crystallography. It may be used in the following key activities. (a)Ab initiophasing,viaPatterson or direct methods. The program may succeed even with structures with up to 6000 non-H atoms in the asymmetric unit, provided that atomic resolution is available, and with data at quasi-atomic resolution (1.4–1.5 Å). (b) Single or multiple isomorphous replacement, single- or multiple-wavelength anomalous diffraction, and single or multiple isomorphous replacement with anomalous scattering techniques. In the first step the program finds the heavy-atom/anomalous scatterer substructure, then automatically uses the above information to phase protein reflections. Phase extension and refinement are performed by electron density modification techniques. (c) Molecular replacement. The orientation and the location of the protein molecules are foundviareciprocal space methods. Phase extension and refinement are performed by electron density modification techniques. All the programs integrated intoIL MILIONEare controlled by means of a user-friendly graphical user interface, which is used to input data and to monitor intermediate and final results by means of real-time updated messages, diagrams and histograms.

scholarly journals Membrane protein structure determination by SAD, SIR, or SIRAS phasing in serial femtosecond crystallography using an iododetergent

2016 ◽  
Vol 113 (46) ◽  
pp. 13039-13044 ◽  
Author(s):  
Takanori Nakane ◽  
Shinya Hanashima ◽  
Mamoru Suzuki ◽  
Haruka Saiki ◽  
Taichi Hayashi ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Structure Determination ◽  
Free Electron ◽  
Heavy Atom ◽  
Anomalous Scattering ◽  
X Ray ◽  
Isomorphous Replacement ◽  
Experimental Phasing ◽  
Serial Femtosecond Crystallography

The 3D structure determination of biological macromolecules by X-ray crystallography suffers from a phase problem: to perform Fourier transformation to calculate real space density maps, both intensities and phases of structure factors are necessary; however, measured diffraction patterns give only intensities. Although serial femtosecond crystallography (SFX) using X-ray free electron lasers (XFELs) has been steadily developed since 2009, experimental phasing still remains challenging. Here, using 7.0-keV (1.771 Å) X-ray pulses from the SPring-8 Angstrom Compact Free Electron Laser (SACLA), iodine single-wavelength anomalous diffraction (SAD), single isomorphous replacement (SIR), and single isomorphous replacement with anomalous scattering (SIRAS) phasing were performed in an SFX regime for a model membrane protein bacteriorhodopsin (bR). The crystals grown in bicelles were derivatized with an iodine-labeled detergent heavy-atom additive 13a (HAD13a), which contains the magic triangle, I3C head group with three iodine atoms. The alkyl tail was essential for binding of the detergent to the surface of bR. Strong anomalous and isomorphous difference signals from HAD13a enabled successful phasing using reflections up to 2.1-Å resolution from only 3,000 and 4,000 indexed images from native and derivative crystals, respectively. When more images were merged, structure solution was possible with data truncated at 3.3-Å resolution, which is the lowest resolution among the reported cases of SFX phasing. Moreover, preliminary SFX experiment showed that HAD13a successfully derivatized the G protein-coupled A2a adenosine receptor crystallized in lipidic cubic phases. These results pave the way for de novo structure determination of membrane proteins, which often diffract poorly, even with the brightest XFEL beams.

scholarly journals Preliminary X-ray diffraction analysis of CfaA, a molecular chaperone essential for the assembly of CFA/I fimbriae of human enterotoxigenicEscherichia coli

2014 ◽  
Vol 70 (2) ◽  
pp. 196-199
Author(s):  
Rui Bao ◽  
Lothar Esser ◽  
Steven Poole ◽  
Annette McVeigh ◽  
Yu-xing Chen ◽  
...  
Keyword(s):  
Tertiary Structure ◽  
Heavy Atom ◽  
Diffusion Method ◽  
Anomalous Scattering ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Colonization Factor ◽  
Isomorphous Replacement ◽  
Periplasmic Chaperone

Understanding of pilus bioassembly in Gram-negative bacteria stems mainly from studies of P pili and type 1 fimbriae of uropathogenicEscherichia coli, which are mediated by the classic chaperone–usher pathway (CUP). However, CFA/I fimbriae, a class 5 fimbria and intestinal colonization factor for enterotoxigenicE. coli(ETEC), are proposed to assembleviathe alternate chaperone pathway (ACP). Both CUP and ACP fimbrial bioassembly pathways require the function of a periplasmic chaperone, but their corresponding proteins share very low similarity in primary sequence. Here, the crystallization of the CFA/I periplasmic chaperone CfaA by the hanging-drop vapor-diffusion method is reported. X-ray diffraction data sets were collected from a native CfaA crystal to 2 Å resolution and to 1.8 and 2.8 Å resolution, respectively, from a lead and a platinum derivative. These crystals displayed the symmetry of space groupC2, with unit-cell parametersa= 103.6,b= 28.68,c= 90.60 Å, β = 119.7°. Initial phases were derived from multiple isomorphous replacement with anomalous scattering experiments using the data from the platinum and lead derivatives. This resulted in an interpretable electron-density map showing one CfaA molecule in an asymmetric unit. Sequence assignments were aided by anomalous signals from the heavy-atom derivatives. Refinement of the atomic model of CfaA is ongoing, which is expected to further understanding of the essential aspects and allowable variations in tertiary structure of the greater family of chaperones involved in chaperone–usher mediated bioassembly.

scholarly journals Alternative phasing method in macromolecular crystallography

2014 ◽  
Vol 70 (a1) ◽  
pp. C342-C342
Author(s):  
Santosh Panjikar ◽  
Daniele de Sanctis
Keyword(s):  
Crystal Structures ◽  
Uv Radiation ◽  
Absorption Edge ◽  
Heavy Atom ◽  
Anomalous Scattering ◽  
Covalent Bonds ◽  
X Ray ◽  
Isomorphous Replacement ◽  
Experimental Phasing ◽  
Radiation Induced

Selenium is the most widely used heavy atom for experimental phasing, either by single anomalous scattering (SAD) or multiple-wavelength anomalous diffraction (MAD) procedures. The use of the single isomorphous replacement (SIR) or single isomorphous replacement with anomalous scattering (SIRAS) phasing procedure with selenomethionine (Mse) containing proteins is not so commonly used, as it requires isomorphous native data. Several non-redundant X-ray diffraction data sets from various Mse derivatised protein crystals were collected at energies far below the absorption edge before and after exposing the crystal to ultraviolet (UV) radiation with 266 nm lasers. A detailed analysis revealed that significant changes in diffracted intensities were induced by ultraviolet irradiation whilst retaining crystal isomorphism. These intensity changes allowed the crystal structures to be solved by the radiation damage-induced phasing (RIP) technique [1]. These can be coupled with the anomalous signal from the dataset collected at the selenium absorption edge to obtain SIRAS phases in a UV-RIPAS phasing experiment [2]. Inspection of the crystal structures and electron-density maps demonstrated that covalent bonds between selenium and carbon at all sites located in the core of the proteins or in a hydrophobic environment were much more susceptible to UV radiation-induced cleavage than other bonds typically present in Mse proteins. The rapid UV radiation-induced bond cleavage opens a reliable new paradigm for phasing at synchrotron [1,2] and at in-house X-ray source [3].

A parallel program usingSHELXDfor quick heavy-atom partial structural solution on high-performance computers

2007 ◽  
Vol 40 (2) ◽  
pp. 387-390 ◽  
Author(s):  
Zheng-Qing Fu ◽  
John Chrzas ◽  
George M. Sheldrick ◽  
John Rose ◽  
Bi-Cheng Wang
Keyword(s):  
High Performance ◽  
National Laboratory ◽  
Heavy Atom ◽  
Argonne National Laboratory ◽  
Scattering Data ◽  
Anomalous Scattering ◽  
Partial Structures ◽  
Multiple Wavelength ◽  
Linux Cluster ◽  
High Performance Computers

A parallel algorithm has been designed forSHELXDto 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.

scholarly journals Introduction to phasing

2010 ◽  
Vol 66 (4) ◽  
pp. 325-338 ◽  
Author(s):  
Garry L. Taylor
Keyword(s):  
Small Molecule ◽  
Heavy Atom ◽  
Protein Crystallography ◽  
Anomalous Scattering ◽  
Phase Information ◽  
X Ray ◽  
Heavy Atoms ◽  
Isomorphous Replacement ◽  
Diffracted Waves ◽  
Complete Protein

When collecting X-ray diffraction data from a crystal, we measure the intensities of the diffracted waves scattered from a series of planes that we can imagine slicing through the crystal in all directions. From these intensities we derive the amplitudes of the scattered waves, but in the experiment we lose the phase information; that is, how we offset these waves when we add them together to reconstruct an image of our molecule. This is generally known as the `phase problem'. We can only derive the phases from some knowledge of the molecular structure. In small-molecule crystallography, some basic assumptions about atomicity give rise to relationships between the amplitudes from which phase information can be extracted. In protein crystallography, theseab initiomethods can only be used in the rare cases in which there are data to at least 1.2 Å resolution. For the majority of cases in protein crystallography phases are derived either by using the atomic coordinates of a structurally similar protein (molecular replacement) or by finding the positions of heavy atoms that are intrinsic to the protein or that have been added (methods such as MIR, MIRAS, SIR, SIRAS, MAD, SAD or combinations of these). The pioneering work of Perutz, Kendrew, Blow, Crick and others developed the methods of isomorphous replacement: adding electron-dense atoms to the protein without disturbing the protein structure. Nowadays, methods from small-molecule crystallography can be used to find the heavy-atom substructure and the phases for the whole protein can be bootstrapped from this prior knowledge. More recently, improved X-ray sources, detectors and software have led to the routine use of anomalous scattering to obtain phase information from either incorporated selenium or intrinsic sulfurs. In the best cases, only a single set of X-ray data (SAD) is required to provide the positions of the anomalous scatters, which together with density-modification procedures can reveal the structure of the complete protein.

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