title: Application of cell-free translation systems to studies of cofactor binding proteins.
abstract: To develop applications of in vitro cell-free translation systems for production and characterization of cofactor binding proteins, we investigate the production of apo- or holo-forms of Flavin Mono Nucleotide (FMN)-binding protein from Desulfovibrio vulgaris (Miyazaki F) and purified them. The redox potential analysis and measurements of UV-, visible, and fluorescent spectra of reconstructed holo-protein showed that the FMN correctly bound to the FMN binding site. On the other hand, contrary to our expectation, we found that the apo-protein formed a dimer structure and the incorporation of the FMN led the conformational alterations of the protein. These studies demonstrate the utility of cell-free translation systems to analyses of cofactor-binding proteins.
authors.affiliation: Department of Applied Chemistry, Faculty of Engineering, Ehime University, Matsuyama 790-8577, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Abe
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Hori
authors.forename: Takeshi
authors.initials: T
authors.lastname: Nakanishi
authors.forename: Fumio
authors.initials: F
authors.lastname: Arisaka
authors.forename: Tomio
authors.initials: T
authors.lastname: Ogasawara
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Sawasaki
authors.forename: Masaya
authors.initials: M
authors.lastname: Kitamura
authors.forename: Yaeta
authors.initials: Y
authors.lastname: Endo
mesh_terms: D001426:Bacterial Proteins / Q000096:biosynthesis / Q000378:metabolism*; D002474:Cell-Free System; D016969:Desulfovibrio vulgaris / Q000378:metabolism*; D005486:Flavin Mononucleotide / Q000378:metabolism*; D011485:Protein Binding; D014176:Protein Biosynthesis; D013997:Time Factors
chemical_list: D001426:Bacterial Proteins; D005486:Flavin Mononucleotide