title: An automated small-scale protein expression and purification screening provides beneficial information for protein production.
abstract: One of the first key steps in structural genomics is high-throughput expression and rapid screening to select highly soluble proteins, the preferred candidates for crystal production. Here we describe the methodology used at the Berkeley Structural Genomics Center (BSGC) for automated parallel expression and small-scale purification of fusion proteins using a 96-well format. Our robotic method includes cell lysis, soluble fraction separation and purification with affinity resins. For detection of His-tagged proteins in the soluble fractions and after affinity resin elution, a dot-blot procedure with an anti-His-antibody is used. The expression level and molecular mass of recombinant proteins are checked by SDS-PAGE. With this approach, we are able to obtain beneficial information to be used for large-scale protein expression and purification.
authors.affiliation: Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
authors.forename: Henry
authors.initials: H
authors.lastname: Nguyen
authors.forename: Bruno
authors.initials: B
authors.lastname: Martinez
authors.forename: Natalia
authors.initials: N
authors.lastname: Oganesyan
authors.forename: Rosalind
authors.initials: R
authors.lastname: Kim
mesh_terms: D001331:Automation; D015870:Gene Expression; D015151:Immunoblotting; D040901:Proteomics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification*; D012371:Robotics; D012995:Solubility
chemical_list: D011994:Recombinant Proteins