title: Biochemical and Structural Analysis of Kinetochore Histone-Fold Complexes.
abstract: The kinetochore connects chromosomes to microtubules during mitosis and therefore plays an essential role in faithful chromosome segregation. It is built at the centromeric region of the chromosome and is comprised of many protein complexes. CENP-S, -T, -W, and -X are kinetochore components with histone-folds. These proteins play important roles in establishment of kinetochore chromatin. Similar to canonical histones, these kinetochore histone-fold proteins form heteromeric complexes (CENP-S/CENP-X complex and CENP-T/CENP-W complex) and bind DNA in sequence independent manner. In addition, they form a CENP-T-W-S-X heterotetrameric complex and bind DNA in a manner that is different from both CENP-S-X and CENP-T-W. To understand how kinetochores form and function it is necessary to characterize the components in detail. Here, we describe our approaches in purification and characterization of the kinetochore histone-fold complexes.
authors.affiliation: Department of Molecular Genetics, National Institute of Genetics and Graduate University for Advanced Studies (SOKENDAI), 1111 Yata, Mishima, Shizuoka, 411-8540, Japan.|Department of Biological Science and Technology, Graduate School of Industrial Science and Technology, Tokyo University of Science, Katsushika-ku, Tokyo, 125-8585, Japan.
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Nishino
authors.affiliation: Department of Molecular Genetics, National Institute of Genetics and Graduate University for Advanced Studies (SOKENDAI), 1111 Yata, Mishima, Shizuoka, 411-8540, Japan. tfukagaw@lab.nig.ac.jp.|Graduate School of Frontier Biosciences, Osaka University, Suita 565-0871, Japan. tfukagaw@lab.nig.ac.jp.
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Fukagawa
mesh_terms: D002503:Centromere / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D020090:Chromosome Segregation; D004247:DNA / Q000378:metabolism; D006657:Histones / Q000737:chemistry* / Q000378:metabolism*; D018386:Kinetochores / Q000737:chemistry* / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D011489:Protein Denaturation; D017510:Protein Folding; D025941:Protein Interaction Mapping; D058849:Protein Refolding; D013329:Structure-Activity Relationship
chemical_list: D002843:Chromatin; D006657:Histones; D046912:Multiprotein Complexes; D004247:DNA
keywords: Centromere; Chromosome segregation; Gel shift assay; Histone-fold; Kinetochore; Protein complex; Protein expression; Protein purification; Protein–DNA interaction; Protein–protein interaction