title: Optimum solubility (OS) screening: an efficient method to optimize buffer conditions for homogeneity and crystallization of proteins.
abstract: One of the most critical steps in the preparation of protein samples for structural studies by X-ray crystallography is to obtain biochemically pure and conformationally homogenous protein samples. Very often, the purified sample does not meet these qualifications and therefore does not crystallize. A screening method, Optimum Solubility Screen, has been developed that consists of two steps. The first step selects a better buffer than that used during purification. 24 different buffers ranging from pH 3 to pH 10 are screened using a vapor-diffusion method and very small amounts of protein. The solubility of the protein is first determined by visual examination using a light microscope and those drops that remain clear after 24 h are further evaluated using dynamic light scattering. If the results from the first step are still not satisfactory, a second step explores a variety of chemical additives in order to improve the monodispersity of the protein sample. In 64% of the cases, crystallization was successful from proteins that had initially shown high levels of aggregation. This screen can be configured to perform in an automated high-throughput mode and can be expanded for additional buffers and additives.
authors.affiliation: Berkeley Structural Genomics Center, Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA.
authors.forename: Jarmila
authors.initials: J
authors.lastname: Jancarik
authors.forename: Ramona
authors.initials: R
authors.lastname: Pufan
authors.forename: Connie
authors.initials: C
authors.lastname: Hong
authors.forename: Sung Hou
authors.initials: SH
authors.lastname: Kim
authors.forename: Rosalind
authors.initials: R
authors.lastname: Kim
mesh_terms: D002021:Buffers; D003460:Crystallization; D018360:Crystallography, X-Ray / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D008027:Light; D008970:Molecular Weight; D009174:Mycoplasma / Q000737:chemistry; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification; D012542:Scattering, Radiation; D012995:Solubility
chemical_list: D002021:Buffers; D011506:Proteins