title: Investigation of the cyclobutane pyrimidine dimer (CPD) photolyase DNA recognition mechanism by NMR analyses.
abstract: The cyclobutane pyrimidine dimer (CPD) is one of the major forms of DNA damage caused by irradiation with ultraviolet (UV) light. CPD photolyases recognize and repair UV-damaged DNA. The DNA recognition mechanism of the CPD photolyase has remained obscure because of a lack of structural information about DNA-CPD photolyase complexes. In order to elucidate the CPD photolyase DNA binding mode, we performed NMR analyses of the DNA-CPD photolyase complex. Based upon results from (31)P NMR measurements, in combination with site-directed mutagenesis, we have demonstrated the orientation of CPD-containing single-stranded DNA (ssDNA) on the CPD photolyase. In addition, chemical shift perturbation analyses, using stable isotope-labeled DNA, revealed that the CPD is buried in a cavity within CPD photolyase. Finally, NMR analyses of a double-stranded DNA (dsDNA)-CPD photolyase complex indicated that the CPD is flipped out of the dsDNA by the enzyme, to gain access to the active site.
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Torizawa
authors.forename: Takumi
authors.initials: T
authors.lastname: Ueda
authors.forename: Seiki
authors.initials: S
authors.lastname: Kuramitsu
authors.forename: Kenichi
authors.initials: K
authors.lastname: Hitomi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Todo
authors.forename: Shigenori
authors.initials: S
authors.lastname: Iwai
authors.forename: Kosuke
authors.initials: K
authors.lastname: Morikawa
authors.forename: Ichio
authors.initials: I
authors.lastname: Shimada
mesh_terms: D001665:Binding Sites; D004247:DNA / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004255:Deoxyribodipyrimidine Photo-Lyase / Q000737:chemistry*; D019281:Dimerization; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008956:Models, Chemical; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011743:Pyrimidines / Q000737:chemistry*; D013053:Spectrophotometry; D020349:Surface Plasmon Resonance; D016963:Thermus thermophilus / Q000378:metabolism; D014466:Ultraviolet Rays
chemical_list: D004277:DNA, Single-Stranded; D011743:Pyrimidines; D004247:DNA; D004255:Deoxyribodipyrimidine Photo-Lyase