title: Data collection with a tailored X-ray beam size at 2.69 Å wavelength (4.6 keV): sulfur SAD phasing of Cdc23(Nterm).
abstract: The capability to reach wavelengths of up to 3.1 Å at the newly established EMBL P13 beamline at PETRA III, the new third-generation synchrotron at DESY in Hamburg, provides the opportunity to explore very long wavelengths to harness the sulfur anomalous signal for phase determination. Data collection at λ = 2.69 Å (4.6 keV) allowed the crystal structure determination by sulfur SAD phasing of Cdc23(Nterm), a subunit of the multimeric anaphase-promoting complex (APC/C). At this energy, Cdc23(Nterm) has an expected Bijvoet ratio〈|Fanom|〉/〈F〉of 2.2%, with 282 residues, including six cysteines and five methionine residues, and two molecules in the asymmetric unit (65.4 kDa; 12 Cys and ten Met residues). Selectively illuminating two separate portions of the same crystal with an X-ray beam of 50 µm in diameter allowed crystal twinning to be overcome. The crystals diffracted to 3.1 Å resolution, with unit-cell parameters a = b = 61.2, c = 151.5 Å, and belonged to space group P43. The refined structure to 3.1 Å resolution has an R factor of 18.7% and an Rfree of 25.9%. This paper reports the structure solution, related methods and a discussion of the instrumentation.
authors.affiliation: EMBL, Notkestrasse 85, 22607 Hamburg, Germany.
authors.forename: Michele
authors.initials: M
authors.lastname: Cianci
authors.affiliation: Structural Biology Unit, Department of Pharmacy, University of Groningen, Antonius Deusinglaan 1, 9713 AV Groningen, The Netherlands.
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Groves
authors.affiliation: Division of Structural Biology, Institute of Cancer Research, Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, England.
authors.forename: David
authors.initials: D
authors.lastname: Barford
authors.affiliation: EMBL, Notkestrasse 85, 22607 Hamburg, Germany.
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Schneider
mesh_terms: D064173:Anaphase-Promoting Complex-Cyclosome / Q000737:chemistry*; D018797:Cell Cycle Proteins / Q000737:chemistry*; D018360:Crystallography, X-Ray / Q000379:methods; D064110:Minichromosome Maintenance Proteins / Q000737:chemistry*; D008958:Models, Molecular; D011487:Protein Conformation; D012568:Schizosaccharomyces / Q000737:chemistry*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry*; D013455:Sulfur / Q000737:chemistry*; D014965:X-Rays
chemical_list: D018797:Cell Cycle Proteins; D029702:Schizosaccharomyces pombe Proteins; C489609:cdc23 protein, S pombe; D013455:Sulfur; D064173:Anaphase-Promoting Complex-Cyclosome; D064110:Minichromosome Maintenance Proteins
keywords: SAD phasing; long wavelength; low resolution; soft X-rays; sulfur