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Molecular-replacement Calculations (molecular-replacement + calculation)
Selected AbstractsCrystallization and preliminary X-ray analysis of Leishmania major glyoxalase IACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2005Antonio Ariza Glyoxalase I (GLO1) is a putative drug target for trypanosomatids, which are pathogenic protozoa that include the causative agents of leishmaniasis. Significant sequence and functional differences between Leishmania major and human GLO1 suggest that it may make a suitable template for rational inhibitor design. L. major GLO1 was crystallized in two forms: the first is extremely disordered and does not diffract, while the second, an orthorhombic form, produces diffraction to 2.0,Å. Molecular-replacement calculations indicate that there are three GLO1 dimers in the asymmetric unit, which take up a helical arrangement with their molecular dyads arranged approximately perpendicular to the c axis. Further analysis of these data are under way. [source] Crystallization and preliminary crystallographic analysis of gallate dioxygenase DesB from Sphingobium sp.ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 11 2009SYK- Gallate dioxygenase (DesB) from Sphingobium sp. SYK-6, which belongs to the type II extradiol dioxygenase family, was purified and crystallized using the hanging-drop vapour-diffusion method. Two crystal forms were obtained. The form I crystal belonged to space group C2, with unit-cell parameters a = 136.2, b = 53.6, c = 55.1,Å, , = 112.8°, and diffracted to 1.6,Å resolution. The form II crystal belonged to space group P21, with unit-cell parameters a = 56.2, b = 64.7, c = 116.1,Å, , = 95.1°, and diffracted to 1.9,Å resolution. A molecular-replacement calculation using LigAB as a search model yielded a satisfactory solution for both crystal forms. [source] MAD phasing using the (Ta6Br12)2+ cluster: a retrospective studyACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2008Oliwia Pasternak The crystal structure of cytokinin-specific binding protein (CSBP) containing four independent molecules with 4 × 155 = 620 residues in the asymmetric unit of the P64 unit cell has been solved by three-wavelength MAD using 1.8,Å resolution data recorded from a crystal derivatized with the dodecabromohexatantalum cation (Ta6Br12)2+. The diffraction data contained a very strong anomalous signal (allowing successful phasing even using peak SAD data alone) despite the fact that the five (Ta6Br12)2+ clusters found in the asymmetric unit have low occupancy (about 0.3). The derivative structure has been successfully refined to R = 0.158, providing interesting details on the geometry of the (Ta6Br12)2+ cluster, its interactions with the protein and on the backsoaking of a cytokinin ligand that was originally part of a CSBP,cytokinin complex in the native crystals used for (Ta6Br12)2+ derivatization. A simulation analysis of the phasing power of the (Ta6Br12)2+ ions at artificially imposed resolution limits shows that it is not possible to resolve the individual Ta atoms if the dmin limit of the data is higher than 2.9,Å. Additionally, for successful Ta identification the (Ta6Br12)2+ complex should be specifically bound and ordered. Good binding at the protein surface is facilitated by the presence of acidic groups, indicating higher pH buffer conditions to be preferable. In addition, the water channels in the crystal should be sufficiently wide (at least 11,Å) to allow free diffusion of the (Ta6Br12)2+ ions on soaking. A retrospective look at the initial molecular-replacement calculations provides interesting insights into how the peculiar packing mode and strong bias of the molecular-replacement-phased electron-density maps had hindered successful solution of the structure by this method. [source] Structure determination of a small protein through a 23-dimensional molecular-replacement searchACTA CRYSTALLOGRAPHICA SECTION D, Issue 4 2003Nicholas M. Glykos The crystal structure of a 4-,-helical bundle protein has been determined by the application of a 23-dimensional molecular-replacement search performed using a stochastic method. The search model for the calculation was a 26-residue-long polyalanine helix amounting to less than 13% of the total number of atoms in the asymmetric unit of the target crystal structure. The crystal structure determination procedure is presented in detail, with emphasis on the molecular-replacement calculations. [source] Crystallization and preliminary X-ray crystallographic analysis of human FAF1 UBX domainACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2010Wonchull Kang Fas-associated factor 1 (FAF1) is a multifunctional pro-apoptotic protein that is involved in Fas-mediated apoptosis, NF-,B signalling and the ubiquitin,proteasome pathway. In the ubiquitin,proteasome pathway, FAF1 binds to the N domain of p97/VCP, a molecular chaperone that acts in complex with the proteasome, through its C-terminal UBX domain and inhibits the proteasomal protein-degradation process. In an effort to elucidate the structural basis of the function of FAF1 in modulating p97/VCP activity related to proteasomal protein degradation, crystallographic analysis of the FAF1 UBX domain and the p97/VCP N domain was initiated. Following the recently reported crystallization of the FAF1 UBX domain bound to the p97/VCP N domain, the unbound FAF1 UBX domain was also crystallized for purposes of structural comparison. X-ray data were collected to 3.00,Å resolution and the crystals belonged to space group F4132, with unit-cell parameters a = b = c = 176.40,Å. The Matthews coefficient and solvent content were estimated to be 3.04,Å3,Da,1 and 59.5%, respectively, assuming that the asymmetric unit contained two molecules of the UBX domain, which was subsequently confirmed by molecular-replacement calculations. [source] Crystallization and preliminary X-ray crystallographic analysis of the N domain of p97/VCP in complex with the UBX domain of FAF1ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2010Hwa Young Shin p97/VCP is a multifunctional AAA+ -family ATPase that is involved in diverse cellular processes. p97/VCP directly interacts with various adaptors for activity in different biochemical contexts. Among these adaptors are p47 and Fas-associated factor 1 (FAF1), which contain a common UBX domain through which they bind to the N domain of p97/VCP. In the ubiquitin,proteasome pathway, p97/VCP acts as a chaperone that presents client proteins to the proteasome for degradation, while FAF1 modulates the process by interacting with ubiquitinated client proteins and also with p97/VCP. In an effort to elucidate the structural details of the interaction between p97/VCP and FAF1, the p97/VCP N domain was crystallized in complex with the FAF1 UBX domain. X-ray data were collected to 2.60,Å resolution and the crystals belonged to space group C2221, with unit-cell parameters a = 58.24, b = 72.81, c = 132.93,Å. The Matthews coefficient and solvent content were estimated to be 2.39,Å3,Da,1 and 48.4%, respectively, assuming that the asymmetric unit contained p97/VCP N domain and FAF1 molecules in a 1:1 ratio, which was subsequently confirmed by molecular-replacement calculations. [source] Purification and crystallization of Cor a 9, a major hazelnut allergenACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2009Feng Guo Hazelnut (Corylus avellana) is one of the food sources that induce allergic reaction in a subpopulation of people with food allergy. The 11S legumin-like seed-storage protein from hazelnut has been identified as one of the major hazelnut allergens and named Cor a 9. In this study, Cor a 9 was extracted from hazelnut kernels using a high-salt solution and was purified by desalting out and FPLC to a highly purified state. Diffraction-quality single crystals were obtained using the hanging-drop vapour-diffusion method. Diffraction data were collected and a structure solution has been obtained by molecular-replacement calculations. Further refinement of the structure is currently in progress. [source] | ||||||||||