title: NMR study of the Z-DNA binding mode and B-Z transition activity of the Zα domain of human ADAR1 when perturbed by mutation on the α3 helix and β-hairpin.
abstract: The Zα domains of human ADAR1 (ZαADAR1) bind to Z-DNA via interaction mediated by the α3-core and β-hairpin. Five residues in the α3 helix and four residues in the β-hairpin play important roles in Zα function, forming direct or water-mediated hydrogen bonds with DNA backbone phosphates or interacting hydrophobically with DNA bases. To understand the roles of these residues during B-Z transition of duplex DNA, we performed NMR experiments on complexes of various ZαADAR1 mutants with a 6-bp DNA duplex at various protein-to-DNA molar ratios. Our study suggests that single mutations at residues K169, N173, or Y177 cause unusual conformational changes in the hydrophobic faces of helices α1, α2, and α3, which dramatically decrease the Z-DNA binding affinity. 1D imino proton spectra and chemical shift perturbation showed that single mutations at residues K170, R174, T191, P192, P193, or W195 slightly affected the Z-DNA binding affinity. A hydrogen exchange study proved that the K170A- and R174A-ZαADAR1 proteins could efficiently change B-DNA to left-handed Z-DNA via an active B-Z transition pathway, whereas the G2·C5 base pair was significantly destabilized compared to wild-type ZαADAR1.
authors.affiliation: Department of Chemistry and Research Institute of Natural Science, Gyeongsang National University, Jinju, Gyeongnam 660-701, Republic of Korea.
authors.forename: Minjee
authors.initials: M
authors.lastname: Jeong
authors.affiliation: Department of Chemistry and Research Institute of Natural Science, Gyeongsang National University, Jinju, Gyeongnam 660-701, Republic of Korea.
authors.forename: Ae-Ree
authors.initials: AR
authors.lastname: Lee
authors.affiliation: Department of Chemistry and Research Institute of Natural Science, Gyeongsang National University, Jinju, Gyeongnam 660-701, Republic of Korea.
authors.forename: Hee-Eun
authors.initials: HE
authors.lastname: Kim
authors.affiliation: Department of Chemistry and Research Institute of Natural Science, Gyeongsang National University, Jinju, Gyeongnam 660-701, Republic of Korea.
authors.forename: Yong-Geun
authors.initials: YG
authors.lastname: Choi
authors.affiliation: Department of Chemistry, KAIST, Daejeon 305-701, Republic of Korea.
authors.forename: Byong-Seok
authors.initials: BS
authors.lastname: Choi
authors.affiliation: Department of Chemistry and Research Institute of Natural Science, Gyeongsang National University, Jinju, Gyeongnam 660-701, Republic of Korea. Electronic address: joonhwa@gnu.ac.kr.
authors.forename: Joon-Hwa
authors.initials: JH
authors.lastname: Lee
mesh_terms: D000243:Adenosine Deaminase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000577:Amides / Q000737:chemistry; D000595:Amino Acid Sequence; D059371:DNA, B-Form / Q000737:chemistry*; D043542:DNA, Z-Form / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D000577:Amides; D059371:DNA, B-Form; D043542:DNA, Z-Form; D000243:Adenosine Deaminase
keywords: ADAR1; B–Z transition; DNA–protein interactions; Hydrogen exchange; NMR; Z-DNA