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Title: Structural characterization of the circadian clock protein complex composed of KaiB and KaiC by inverse contrast-matching small-angle neutron scattering
Authors: Sugiyama, Masaaki
Yagi, Hirokazu
Ishii, Kentaro
Porcar, Lionel
Martel, Anne
Oyama, Katsuaki
Noda, Masanori
Yunoki, Yasuhiro
Murakami, Reiko
Inoue, Rintaro  kyouindb  KAKEN_id
Sato, Nobuhiro  kyouindb  KAKEN_id
Oba, Yojiro
Terauchi, Kazuki
Uchiyama, Susumu
Kato, Koichi
Author's alias: 杉山, 正明
Keywords: Mass spectrometry
Structural biology
Issue Date: 18-Oct-2016
Publisher: Springer Nature
Journal title: Scientific Reports
Volume: 6
Thesis number: 35567
Abstract: The molecular machinery of the cyanobacterial circadian clock consists of three proteins: KaiA, KaiB, and KaiC. Through interactions among the three Kai proteins, the phosphorylation states of KaiC generate circadian oscillations in vitro in the presence of ATP. Here, we characterized the complex formation between KaiB and KaiC using a phospho-mimicking mutant of KaiC, which had an aspartate substitution at the Ser431 phosphorylation site and exhibited optimal binding to KaiB. Mass-spectrometric titration data showed that the proteins formed a complex exclusively in a 6:6 stoichiometry, indicating that KaiB bound to the KaiC hexamer with strong positive cooperativity. The inverse contrast-matching technique of small-angle neutron scattering enabled selective observation of KaiB in complex with the KaiC mutant with partial deuteration. It revealed a disk-shaped arrangement of the KaiB subunits on the outer surface of the KaiC C1 ring, which also serves as the interaction site for SasA, a histidine kinase that operates as a clock-output protein in the regulation of circadian transcription. These data suggest that cooperatively binding KaiB competes with SasA with respect to interaction with KaiC, thereby promoting the synergistic release of this clock-output protein from the circadian oscillator complex.
Rights: This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
URI: http://hdl.handle.net/2433/219650
DOI(Published Version): 10.1038/srep35567
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