title: Getting the most out of X-ray home sources.
abstract: The structures of a 14 kDa phospholipase, an 18 kDa proteinase inhibitor and a novel glycoside hydrolase with molecular weight 60 kDa were solved using the SAD technique and the effects of the amount of anomalous signal, completeness and redundancy of data on heavy-atom substructure determination, phasing and model building were analyzed. All diffraction data sets were collected on a Cu-anode X-ray home source. The structure of the phospholipase was obtained using the anomalous scattering contribution from its 16 S atoms. Three-dimensional models for the other two macromolecules were obtained using the anomalous contribution of I atoms rapidly incorporated into the crystal through the quick cryo-soaking method of derivatization. These results were used to discuss the application of sulfur- and iodine-SAD approaches in combination with X-ray home sources for high-throughput protein crystal structure solution. The estimates of the anomalous signal from S atoms in the gene products of four genomes are given and the prospects for increasing the anomalous contribution using longer wavelengths (e.g. from a chromium home source) and quick cryo-soaking derivatization are discussed. The possibility of rapidly preparing tangible home-source isomorphous derivatives suggests that this approach might become a valuable tool in the future of post-genomic projects.
authors.affiliation: Institute of Physics of São Carlos, University of São Paulo, Avenida Trabalhador Sãocarlense 400, CEP 13560-970, São Carlos, SP, Brazil.
authors.forename: R A P
authors.initials: RA
authors.lastname: Nagem
authors.forename: A L B
authors.initials: AL
authors.lastname: Ambrosio
authors.forename: A L
authors.initials: AL
authors.lastname: Rojas
authors.forename: M V A S
authors.initials: MV
authors.lastname: Navarro
authors.forename: A M
authors.initials: AM
authors.lastname: Golubev
authors.forename: R C
authors.initials: RC
authors.lastname: Garratt
authors.forename: I
authors.initials: I
authors.lastname: Polikarpov
mesh_terms: D001230:Aspergillus / Q000201:enzymology; D036481:Bauhinia / Q000737:chemistry; D003460:Crystallization; D018360:Crystallography, X-Ray / Q000379:methods*; D016680:Genome, Bacterial; D006026:Glycoside Hydrolases / Q000737:chemistry*; D008958:Models, Molecular; D010741:Phospholipases A / Q000737:chemistry*; D011480:Protease Inhibitors / Q000737:chemistry*; D013455:Sulfur / Q000737:chemistry; D014965:X-Rays
chemical_list: D011480:Protease Inhibitors; D013455:Sulfur; D010741:Phospholipases A; D006026:Glycoside Hydrolases