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Lattice Formation (lattice + formation)
Selected AbstractsMaternal-effect gene Ces5/Ooep/Moep19/Floped is essential for oocyte cytoplasmic lattice formation and embryonic development at the maternal-zygotic stage transitionGENES TO CELLS, Issue 8 2010Fumi Tashiro In a search for genes specifically expressed in mouse embryonic stem cells, we identified one we called Ces5. We found that it corresponded to the Ooep gene, which was recently reported to be expressed specifically in oocytes. Mouse Ces5/Ooep, also called Moep19 or Floped, encoded a 164-amino acid protein, which was detected in the cytoplasm of developing and mature oocytes and in embryos throughout the preimplantation period. To examine its function, we carried out targeted disruption of this gene. The Ces5/Ooep -null mice were grossly normal, but the females were infertile. Although the ovaries and ovulation appeared normal, the embryos from Ces5/Ooep -null females mated with wild-type males showed developmental arrest at the two- or four-cell stage. In addition, their first cleavage was considerably delayed and often asymmetrical. Thus, Ces5/Ooep is a maternal-effect gene. By electron microscopy, we found that the eggs from Ces5/Ooep -null females lacked oocyte cytoplasmic lattices (CPLs), which have long been predicted to function as a storage form for components that are maternally contributed to the early embryo. Further analysis showed that CES5/OOEP was directly associated with the CPLs. These results indicate that CES5/OOEP is an essential component of the CPLs and is required for embryonic development at the maternal-zygotic stage transition. [source] A synergistic approach to protein crystallization: Combination of a fixed-arm carrier with surface entropy reductionPROTEIN SCIENCE, Issue 5 2010Andrea F. Moon Abstract Protein crystallographers are often confronted with recalcitrant proteins not readily crystallizable, or which crystallize in problematic forms. A variety of techniques have been used to surmount such obstacles: crystallization using carrier proteins or antibody complexes, chemical modification, surface entropy reduction, proteolytic digestion, and additive screening. Here we present a synergistic approach for successful crystallization of proteins that do not form diffraction quality crystals using conventional methods. This approach combines favorable aspects of carrier-driven crystallization with surface entropy reduction. We have generated a series of maltose binding protein (MBP) fusion constructs containing different surface mutations designed to reduce surface entropy and encourage crystal lattice formation. The MBP advantageously increases protein expression and solubility, and provides a streamlined purification protocol. Using this technique, we have successfully solved the structures of three unrelated proteins that were previously unattainable. This crystallization technique represents a valuable rescue strategy for protein structure solution when conventional methods fail. [source] Analysis of multiple crystal forms of Bacillus subtilis BacB suggests a role for a metal ion as a nucleant for crystallizationACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2010M. Rajavel Bacillus subtilis BacB is an oxidase that is involved in the production of the antibiotic bacilysin. This protein contains two double-stranded ,-helix (cupin) domains fused in a compact arrangement. BacB crystallizes in three crystal forms under similar crystallization conditions. An interesting observation was that a slight perturbation of the crystallization droplet resulted in the nucleation of a different crystal form. An X-ray absorption scan of BacB suggested the presence of cobalt and iron in the crystal. Here, a comparative analysis of the different crystal forms of BacB is presented in an effort to identify the basis for the different lattices. It is noted that metal ions mediating interactions across the asymmetric unit dominate the different packing arrangements. Furthermore, a normalized B -factor analysis of all the crystal structures suggests that the solvent-exposed metal ions decrease the flexibility of a loop segment, perhaps influencing the choice of crystal form. The residues coordinating the surface metal ion are similar in the triclinic and monoclinic crystal forms. The coordinating ligands for the corresponding metal ion in the tetragonal crystal form are different, leading to a tighter packing arrangement. Although BacB is a monomer in solution, a dimer of BacB serves as a template on which higher order symmetrical arrangements are formed. The different crystal forms of BacB thus provide experimental evidence for metal-ion-mediated lattice formation and crystal packing. [source] Dramatic improvement of crystal quality for low-temperature-grown rabbit muscle aldolaseACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2010HaJeung Park Rabbit muscle aldolase (RMA) was crystallized in complex with the low-complexity domain (LC4) of sorting nexin 9. Monoclinic crystals were obtained at room temperature that displayed large mosaicity and poor X-ray diffraction. However, orthorhombic RMA,LC4 crystals grown at 277,K under similar conditions exhibited low mosaicity, allowing data collection to 2.2,Å Bragg spacing and structure determination. It was concluded that the improvement of crystal quality as indicated by the higher resolution of the new RMA,LC4 complex crystals was a consequence of the introduction of new lattice contacts at lower temperature. The lattice contacts corresponded to an increased number of interactions between high-entropy side chains that mitigate the lattice strain incurred upon cryocooling and accompanying mosaic spread increases. The thermodynamically unfavorable immobilization of high-entropy side chains used in lattice formation was compensated by an entropic increase in the bulk-solvent content owing to the greater solvent content of the crystal lattice. [source] An improved strategy for the crystallization of Leishmania mexicana pyruvate kinaseACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2010Hugh P. Morgan The inclusion of novel small molecules in crystallization experiments has provided very encouraging results and this method is now emerging as a promising alternative strategy for crystallizing `problematic' biological macromolecules. These small molecules have the ability to promote lattice formation through stabilizing intermolecular interactions in protein crystals. Here, the use of 1,3,6,8-pyrenetetrasulfonic acid (PTS), which provides a helpful intermolecular bridge between Leishmania mexicana PYK (LmPYK) macromolecules in the crystal, is reported, resulting in the rapid formation of a more stable crystal lattice at neutral pH and greatly improved X-ray diffraction results. The refined structure of the LmPYK,PTS complex revealed the negatively charged PTS molecule to be stacked between positively charged (surface-exposed) arginine side chains from neighbouring LmPYK molecules in the crystal lattice. [source] | ||||||||||