Structure of tetragonal hen egg-white lysozyme at 0.94 A from crystals grown by the counter-diffusion method.
Very high quality crystals of tetragonal hen egg-white lysozyme were grown in the Advanced Protein Crystallization Facility (APCF) on board the Space Shuttle using a modified free-interface diffusion (FID) reactor designed ad hoc to have a longer diffusion path. This design allows the performance of true counter-diffusion experiments. Crystals were obtained under the classical chemical conditions defined 50 y ago with NaCl as a crystallizing agent and acetate pH 4.5 as a buffer. Counter-diffusion crystallization allows a "physical" instead of chemical optimization of growth conditions: indeed, this method screens for the best supersaturation conditions in a single trial and yields crystals of very high quality. A complete diffraction data set was collected at atomic resolution from one of these crystals using synchrotron radiation at the DESY-EMBL beamlines. The overall R(merge) on intensities in the resolution range 31-0.94 A was 5.2% and the data were 98.9% complete. Refinement was carried out with the programs CNS and SHELX97 to a final crystallographic R factor of 12.26% for 72 390 reflections. A mean standard uncertainty in the atomic positions of 0.024 A was estimated from inversion of blocked least-squares matrices. 22 side chains show alternate conformations and the loop 59-75 adopts in the same crystal packing two conformations that were observed for either triclinic or tetragonal lysozyme in previous high-resolution studies. In addition to 255 water molecules, the crystallizing agent (one hexacoordinated sodium ion and five chloride anions) participates in the ordered lysozyme hydration shell.
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UPR 9002, Département Mécanismes et Molécules de la Synthèse Protéique et Cristallogenèse, Institut de Biologie Moléculaire et Cellulaire, 15 Rue René Descartes, F-67084 Strasbourg CEDEX, France.
Otálora
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Gavira
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Vidal
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Giegé
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García-Ruiz
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D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D000838:Anions / Q000737:chemistry; D001665:Binding Sites; D002412:Cations / Q000737:chemistry; D002645:Chickens; D003460:Crystallization; D018360:Crystallography, X-Ray; D004529:Egg White / Q000032:analysis; D008958:Models, Molecular; D009113:Muramidase / Q000737:chemistry*; D011487:Protein Conformation; D017434:Protein Structure, Tertiary
D000596:Amino Acids; D000838:Anions; D002412:Cations; C421864:hen egg lysozyme; D009113:Muramidase