Protein crystal structure solution by fast incorporation of negatively and positively charged anomalous scatterers

The preparation of derivatives by the traditional methods of soaking is one of the most time-consuming steps in protein crystal structure solution by X-ray diffraction techniques. The `quick cryosoaking' procedure for derivatization with halides (monovalent anions) offers the possibility of significantly speeding up this process [Dauter et al. (2000), Acta Cryst. D56, 232-237]. In the present work, an extension of this technique is proposed and the use of two different classes of compounds (monovalent and polyvalent cations) that can be successfully utilized in the quick cryosoaking procedure for the derivatization and phasing of protein crystals is described. This approach has been tested on hen egg-white lysozyme and has been successfully used to solve the structure of a novel trypsin inhibitor. The possibility of using cations in the fast cryosoaking procedure gives additional flexibility in the process of derivatization and increases the chances of success in phase determination. This method can be applied to high-throughput crystallographic projects.577996100

Crystallization and synchrotron X-ray diffraction studies of human interleukin-22

Human interleukin-22, a novel member of the cytokine family, has been crystallized in hanging drops using the vapour-diffusion technique. Preliminary X-ray diffraction experiments using synchrotron radiation reveal that the protein crystallizes in space group P2(1)2(1)2(1), with unit-cell parameters a = 55.44, b = 61.62, c = 73.43 Angstrom, and diffracts beyond 2.00 Angstrom resolution.58352953

Crystallization and preliminary X-ray study of haem-binding protein from the bloodsucking insect Rhodnius prolixus

Rhodnius haem-binding protein (RHBP) from the bloodsucking insect Rhodnius prolixus, a 15 kDa protein, has been crystallized using polyethylene glycol as a precipitant. X-ray diffraction data have been collected at a synchrotron source. The crystals belong to the space group P4(1(3))2(1)2, with unit-cell parameters a = b = 64.98, c = 210.68 Angstrom, and diffract beyond 2.6 Angstrom resolution.57686086

Crystallization and preliminary X-ray diffraction analysis of a novel trypsin inhibitor from seeds of Copaifera langsdorffii

A novel trypsin inhibitor isolated from seeds of Copaifera langsdorffii was purified to homogeneity and crystallized. Crystals suitable for X-ray analysis were grown using the hanging-drop vapour-diffusion method at 291 K in sodium acetate buffer at pH values near 4.3 using PEG 4000 as precipitant. The crystals presented symmetry compatible with the space group P4(1)2(1)2 or P4(3)2(1)2, with unit-cell parameters a = b = 58.71, c = 93.75 Angstrom, and diffracted to 1.83 Angstrom resolution at the synchrotron source.5791316131

Direct way to anomalous scatterers

The first step in solving macromolecular crystal structures by multi- or single-wavelength anomalous diffraction methods is the location of the anomalous scatterers. This can be done by direct methods, using either Bijvoet differences within the single data set, or anomalous scattering amplitudes estimated from measurements at several wavelengths. The calculations suggest that Bijvoet differences are equally successful for this purpose as anomalous amplitudes, F(A), which theoretically should be more suitable. The calculation of F(A) values is susceptible to the accumulation of errors contained in the individual intensity measurements at several wavelengths and in the inaccurate estimation of the anomalous atomic scattering corrections, f' and f'. Direct methods often give better results at resolution lower than the full extent of the diffraction data limit. This may be attributed to the enhanced accuracy of measurements of the strong, low resolution reflections and to more effective phase refinement and propagation through Sigma(2) relations.o TEXTO COMPLETO DESTE ARTIGO, ESTARÁ DISPONÍVEL À PARTIR DE AGOSTO DE 2015.2171269470

Three-dimensional structure of an unusual Kunitz (STI) type trypsin inhibitor from Copaifera langsdorffii

The crystallographic structure of a novel trypsin inhibitor (CTI) from Copaifera langsdorffti is reported. The structure was solved by MIRAS procedure and refined to a crystallographic residual of 17.3% (R-free = 20.3%) at 1.8 Angstrom resolution. Two isomorphous derivatives were obtained by quick cryo-soaking approach. CTI is the first structure of a member of Kunitz (STI) family formed by two noncovalently bound polypeptide chains and only one disulfide bridge. A standard Kunitz-type inhibitor has a single polypeptide chain and two disulfide bridges. Structural features granting CTI high inhibitory activity are discussed. (C) 2004 Elsevier SAS. All rights reserved.86316717

Crystal structure of recombinant human interleukin-22

Interleukin-22 (IL-10-related T cell-derived inducible factor/IL-TIF/IL-22) is a novel cytokine belonging to the IL-10 family. Recombinant human IL-22 (hIL-22) was found to activate the signal transducers and activators of transcription factors I and 3 as well as acute phase reactants in several hepatoma cell lines, suggesting its involvement in the inflammatory response. The crystallographic structure of recombinant hIL-22 has been solved at 2.0 Angstrom resolution using the SIRAS method. Contrary to IL-10, the hIL-22 dimer does not present an interpenetration of the secondary-structure elements belonging to the two distinct polypeptide chains but results from interface interactions between monomers. Structural differences between these two cytokines, revealed by the crystallographic studies, clearly indicate that, while a homodimer of IL-10 is required for signaling, hIL-22 most probably interacts with its receptor as a monomer.1081051106

Crystal structure of recombinant human interleukin-22

Interleukin-22 (IL-10-related T cell-derived inducible factor/IL-TIF/IL-22) is a novel cytokine belonging to the IL-10 family. Recombinant human IL-22 (hIL-22) was found to activate the signal transducers and activators of transcription factors I and 3 as well as acute phase reactants in several hepatoma cell lines, suggesting its involvement in the inflammatory response. The crystallographic structure of recombinant hIL-22 has been solved at 2.0 Angstrom resolution using the SIRAS method. Contrary to IL-10, the hIL-22 dimer does not present an interpenetration of the secondary-structure elements belonging to the two distinct polypeptide chains but results from interface interactions between monomers. Structural differences between these two cytokines, revealed by the crystallographic studies, clearly indicate that, while a homodimer of IL-10 is required for signaling, hIL-22 most probably interacts with its receptor as a monomer

Crystallization and synchrotron X-ray diffraction studies of human interleukin-22

Human interleukin-22, a novel member of the cytokine family, has been crystallized in hanging drops using the vapour-diffusion technique. Preliminary X-ray diffraction experiments using synchrotron radiation reveal that the protein crystallizes in space group P2(1)2(1)2(1), with unit-cell parameters a = 55.44, b = 61.62, c = 73.43 Angstrom, and diffracts beyond 2.00 Angstrom resolution