title: Conformational transition of DnaA protein by ATP: structural analysis of DnaA protein, the initiator of Escherichia coli chromosome replication.
abstract: DnaA protein binds to the chromosomal origin (oriC) to initiate DNA replication. We developed an efficient system for purification of DnaA protein which will facilitate physicochemical analysis of the protein. The yield of DnaA protein was increased at least 6-fold compared to an available method being used, and over 22 mg of the protein were obtained from only 100 g of cells. DnaA protein purified by this procedure showed an indistinguishable affinity for ATP, and activity for in vitro replication of oriC plasmid. The process of denaturation of DnaA protein, which was blocked by ATP, was monitored by intrinsic fluorescence and circular dichroism. Analysis of circular dichroism revealed that DnaA protein is rich in alpha-helices, and that ATP-binding leads to a significant transition of protein conformation in that the content of alpha-helices is decreased. This is the first evidence indicating that ATP-binding profoundly affects conformation of DnaA protein.
authors.affiliation: Department of Microbiology, Kyushu University Faculty of Pharmaceutical Sciences, Fukuoka, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Kubota
authors.forename: T
authors.initials: T
authors.lastname: Katayama
authors.forename: Y
authors.initials: Y
authors.lastname: Ito
authors.forename: T
authors.initials: T
authors.lastname: Mizushima
authors.forename: K
authors.initials: K
authors.lastname: Sekimizu
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry* / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005453:Fluorescence; D011485:Protein Binding; D017433:Protein Structure, Secondary
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; D000255:Adenosine Triphosphate