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Hen Egg-white Lysozyme Crystals (hen + egg-white_lysozyme_crystal)
Selected AbstractsPhase transition of triclinic hen egg-white lysozyme crystal associated with sodium bindingACTA CRYSTALLOGRAPHICA SECTION D, Issue 4 2004Kazuaki Harata A triclinic crystal of hen egg-white lysozyme obtained from a D2O solution at 313,K was transformed into a new triclinic crystal by slow release of solvent under a temperature-regulated nitrogen-gas stream. The progress of the transition was monitored by X-ray diffraction. The transition started with the appearance of strong diffuse streaks. The diffraction spots gradually fused and faded with the emergence of diffraction from the new lattice; the scattering power of the crystal fell to a resolution of 1.5,Å from the initial 0.9,Å resolution. At the end of the transition, the diffuse streaks disappeared and the scattering power recovered to 1.1,Å resolution. The transformed crystal contained two independent molecules and the solvent content had decreased to 18% from the 32% solvent content of the native crystal. The structure was determined at 1.1,Å resolution and compared with the native structure refined at the same resolution. The backbone structures of the two molecules in the transformed crystal were superimposed on the native structure with root-mean-square deviations of 0.71 and 0.96,Å. A prominent structural difference was observed in the loop region of residues Ser60,Leu75. In the native crystal, a water molecule located at the centre of this helical loop forms hydrogen bonds to main-chain peptide groups. In the transformed crystal, this water molecule is replaced by a sodium ion with octahedral coordination that involves water molecules and a nitrate ion. The peptide group connecting Arg73 and Asn74 is rotated by 180° so that the CO group of Arg73 can coordinate to the sodium ion. The change in the X-ray diffraction pattern during the phase transition suggests that the transition proceeds at the microcrystal level. A mechanism is proposed for the crystal transformation. [source] Macromolecular crystallography at high pressure with pneumatic diamond anvil cells handled by a six-axis robotic armJOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 4 2010Eric Girard A new pneumatic diamond anvil cell has been constructed, generating continuous pressure and temperature variations in the range 0,2.5,GPa and 293,393,K. The cell is designed mainly for high-pressure macromolecular crystallography and should facilitate pressure and temperature annealing of the sample. The analysis is reported of several diffraction data sets of tetragonal hen egg-white lysozyme crystals loaded either in the new cell or in a currently used membrane-based cell. These experiments were performed on beamline FIP-BM30A at the ESRF, Grenoble, a macromolecular crystallography beamline on a bending magnet. Cells were handled and automatically centred by a six-axis robotic arm that was used as a goniometer for data collection by the oscillation method. [source] Identification of dislocations in large tetragonal hen egg-white lysozyme crystals by synchrotron white-beam topographyJOURNAL OF SYNCHROTRON RADIATION, Issue 6 2003M. Tachibana Large tetragonal hen egg-white (HEW) lysozyme crystals have been grown by a salt concentration-gradient method. The grown crystals, of thicknesses greater than 1.5,mm, were observed by means of X-ray topography using white-beam synchrotron radiation. Line contrasts clearly appeared on the Laue topographs. Extinction of the line images was observed in specific reflections. These results mean that the observed lines correspond to dislocation images. From the extinction criterion it is shown that the predominant dislocations are of screw character with ,110, Burgers vectors. In addition, dislocation loops with [001] Burgers vectors have been found in a tetragonal HEW lysozyme crystal including some cracks. These results are discussed in the light of dislocation elastic energy and slip systems in the crystals. [source] The effect of protein,precipitant interfaces and applied shear on the nucleation and growth of lysozyme crystalsACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2009Nuno M. Reis This paper is concerned with the effect of protein,precipitant interfaces and externally applied shear on the nucleation and growth kinetics of hen egg-white lysozyme crystals. The early stages of microbatch crystallization of lysozyme were explored using both optical and confocal fluorescence microscopy imaging. Initially, an antisolvent (precipitant) was added to a protein drop and the optical development of the protein,precipitant interface was followed with time. In the presence of the water-soluble polymer poly(ethylene glycol) (PEG) a sharp interface was observed to form immediately within the drop, giving an initial clear separation between the lighter protein solution and the heavier precipitant. This interface subsequently became unstable and quickly developed within a few seconds into several unstable `fingers' that represented regions of high concentration-gradient interfaces. Confocal microscopy demonstrated that the subsequent nucleation of protein crystals occurred preferentially in the region of these interfaces. Additional experiments using an optical shearing system demonstrated that oscillatory shear significantly decreased nucleation rates whilst extending the growth period of the lysozyme crystals. The experimental observations relating to both nucleation and growth have relevance in developing efficient and reliable protocols for general crystallization procedures and the controlled crystallization of single large high-quality protein crystals for use in X-ray crystallography. [source] The interdependence of wavelength, redundancy and dose in sulfur SAD experimentsACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2008Michele Cianci In the last decade, the popularity of sulfur SAD anomalous dispersion experiments has spread rapidly among synchrotron users as a quick and streamlined way of solving the phase problem in macromolecular crystallography. On beamline 10 at SRS (Daresbury Laboratory, UK), a versatile design has allowed test data sets to be collected at six wavelengths between 0.979 and 2.290,Å in order to evaluate the importance and the interdependence of experimental variables such as the Bijvoet ratio, wavelength, resolution limit, data redundancy and absorbed X-ray dose in the sample per data set. All the samples used in the experiments were high-quality hen egg-white lysozyme crystals. X-radiation damage was found to affect disulfide bridges after the crystals had been given a total dose of 0.20 × 107,Gy. However, with such a total dose, it was still possible in all cases to find a strategy to collect data sets to determine the sulfur substructure and produce good-quality phases by choosing an optimum combination of wavelength, exposure time and redundancy. A ,|,ano|/,(,ano), greater than 1.5 for all resolution shells was a necessary requirement for successful sulfur SAD substructure location. Provided this is achieved, it seems possible to find an optimum compromise between wavelength, redundancy and dose to provide phasing information. The choice of the wavelength should then follow the sample composition and the diffracting properties of the crystal. For strongly diffracting crystals, wavelengths equal or shorter than 1.540,Å can be selected to capture the available data (provided the Bijvoet ratio is reasonable), while a longer wavelength, to gain as high a Bijvoet ratio as possible, must be used for more weakly diffracting crystals. These results suggest that an approach to a sulfur SAD experiment based on a complete description of the crystal system and the instrument for data collection is useful. [source] Structure determination of a novel protein by sulfur SAD using chromium radiation in combination with a new crystal-mounting methodACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2005Yu Kitago A novel and easy crystal-mounting technique was developed for the sulfur SAD method using Cr,K, radiation (2.29,Å). Using this technique, the cryo-buffer and cryoloop around the protein crystal can be removed before data collection in order to eliminate their X-ray absorption. The superiority and reproducibility of the data sets with this mounting technique were demonstrated using tetragonal hen egg-white lysozyme crystals. The structure of a novel protein, PH1109, from Pyrococcus horikoshii OT3 was solved using this technique. At the wavelength of Cr,K, radiation, the anomalous signal ,|,F|,/,|F|, of PH1109 is expected to be 1.72% as this protein of 144 residues includes four methionines and two cysteines. Sulfur SAD phasing was performed using SHELXD and SHELXE. In the case of the data set obtained using this novel crystal-mounting technique, 54.9% of all residues were built with side chains automatically by RESOLVE. On the other hand, only 16.0% were built with side chains for the data set collected using the standard cryoloop. These results indicated that this crystal-mounting technique was superior to the standard loop-mounting method for the measurement of small anomalous differences at longer wavelength and yielded better results in sulfur-substructure solution and initial phasing. The present study demonstrates that the sulfur SAD method with a chromium source becomes enhanced and more practical for macromolecular structure determination using the new crystal-mounting technique. [source] | ||||||||||