title: Heme binds to an intrinsically disordered region of Bach2 and alters its conformation.
abstract: The transcriptional repressor Bach2 regulates humoral and cellular immunity, including antibody class switching. It possesses a basic leucine zipper domain that mediates DNA binding. Heme inhibits the DNA-binding activity of Bach2 in vitro and induces the degradation of Bach2 in B cells. However, the structural basis of the heme-Bach2 interaction has not been identified. Spectroscopic analyses revealed that Bach2(331-520) is the heme-binding domain, as it includes three Cys-Pro motifs known to be important for heme binding. Heme-titration experiments demonstrated the presence of 5- and 6-coordinated heme-binding modes. Circular dichroism measurements indicated that Bach2(331-520) exists mostly in a random-coil conformation. However, dynamic light scattering analyses showed that, upon heme binding to Bach2(331-520), this region becomes denatured at a lower temperature, as compared with unbound Bach2(331-520). In addition, small-angle X-ray scattering and chemical modification analyses revealed that heme binding induces conformational alterations within the unstructured region. A GAL4-based luciferase assay in 293T cells showed that heme alters the protein interactions mediated by Bach2(331-520). These observations suggested that the unstructured region of Bach2 is important for heme binding, and consequently for its functional regulation.
authors.affiliation: Department of Biochemistry, Tohoku University Graduate School of Medicine, Sendai, Japan.
authors.forename: Miki
authors.initials: M
authors.lastname: Watanabe-Matsui
authors.affiliation: Rigaku Corporation, Akishima, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Matsumoto
authors.affiliation: Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, Sendai, Japan.
authors.forename: Toshitaka
authors.initials: T
authors.lastname: Matsui
authors.affiliation: Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, Sendai, Japan.
authors.forename: Masao
authors.initials: M
authors.lastname: Ikeda-Saito
authors.affiliation: Department of Biochemistry, Tohoku University Graduate School of Medicine, Sendai, Japan.
authors.forename: Akihiko
authors.initials: A
authors.lastname: Muto
authors.affiliation: Division of Biomedical Measurements and Diagnostics, Tohoku University Graduate School of Biomedical Engineering, Sendai, Japan; Division of Structural and Synthetic Biology, RIKEN Yokohama Institute, Yokohama, Japan. Electronic address: kmura@bme.tohoku.ac.jp.
authors.forename: Kazutaka
authors.initials: K
authors.lastname: Murayama
authors.affiliation: Department of Biochemistry, Tohoku University Graduate School of Medicine, Sendai, Japan; Center for Regulatory Epigenome and Diseases, Tohoku University Graduate School of Medicine, Sendai, Japan; CREST, Japan Science and Technology Agency, Sendai, Japan. Electronic address: igarashi@med.tohoku.ac.jp.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Igarashi
mesh_terms: D050976:Basic-Leucine Zipper Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D006418:Heme / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016350:Leucine Zippers; D011485:Protein Binding; D017434:Protein Structure, Tertiary
chemical_list: C491123:BACH2 protein, human; D050976:Basic-Leucine Zipper Transcription Factors; D006418:Heme
keywords: Chemical modification; Conformation change; Heme binding; Intrinsically disordered protein; Small-angle X-ray scattering