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Improved Methods for Preparing the Telomere Tethering Complex Bqt1–Bqt2 for Structural Studies

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Abstract

In eukaryotes, chromosome ends (telomeres) are tethered to the inner nuclear membrane. During the early stages of meiosis, telomeres move along the nuclear membrane and gather near the spindle-pole body, resulting in a bouquet-like arrangement of chromosomes. This chromosomal configuration appears to be widely conserved among eukaryotes, and is assumed to play an important role in the normal progression of meiosis, by mediating the proper pairing of homologous chromosomes. In fission yeast, the Bqt1–Bqt2 protein complex plays a key role in tethering the telomere to the inner nuclear membrane. However, the structural details of the complex required to clarify how telomeres are gathered near the spindle-pole body remain enigmatic. Previously, we devised a preparation procedure for the Schizosaccharomyces japonicus Bqt1–Bqt2 complex, in which a SUMO tag was fused to the N-terminus of the Bqt1 protein. This allowed us to purify the Bqt1–Bqt2 complex from the soluble fraction. In the present study, we found that a maltose-binding protein homolog, Athe_0614, served as a better fusion partner than the SUMO protein, resulting in the marked increase in the solubility of the Bqt1–Bqt2 complex. The Athe_0614 fusion partner may open up new avenues for X-ray crystallographic analyses of the structure of the Bqt1–Bqt2 complex.

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Funding

This study was funded by JSPS KAKENHI Grant Numbers: JP19K12328, JP24570138, JP26116521 and JP16H01316 (to W.K.); JP18H05534 and JP17H01408 (to H.K.); JP18H05528 (to T.H.); JP18H05533 (to Y.H.). W.K. was also supported by the Science Research Promotion Fund from the Japan Private School Promotion Foundation, and by the Priority Research Funding from Meisei University.

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Authors and Affiliations

  1. Department of Chemistry, Graduate School of Science and Engineering, Meisei University, 2-1-1 Hodokubo, Hino-shi, Tokyo, 191-8506, Japan

    Hidetaka Yuzurihara, Yuuki Aizawa, Mika Saotome & Wataru Kagawa

  2. Division of Cancer Biology, The Cancer Institute of JFCR, 3-8-31 Ariake, Koto-ku, Tokyo, 135-8550, Japan

    Yuichi Ichikawa

  3. Animal Waste Management and Environment Division, National Institute of Livestock and Grassland Science, 2 Ikenodai, Tsukuba, Ibaraki, 305-0901, Japan

    Hiroshi Yokoyama

  4. Advanced ICT Research Institute Kobe, National Institute of Information and Communications Technology, 588-2 Iwaoka, Iwaoka-cho, Nishi-ku, Kobe, 651-2492, Japan

    Yuji Chikashige, Tokuko Haraguchi & Yasushi Hiraoka

  5. Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, 565-0871, Japan

    Tokuko Haraguchi & Yasushi Hiraoka

  6. Laboratory of Chromatin Structure and Function, Institute for Quantitative Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Bunkyo-ku, Tokyo, 113-0032, Japan

    Hitoshi Kurumizaka

Authors
  1. Hidetaka Yuzurihara
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  2. Yuuki Aizawa
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  3. Mika Saotome
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  4. Yuichi Ichikawa
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  5. Hiroshi Yokoyama
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  6. Yuji Chikashige
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  7. Tokuko Haraguchi
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  8. Yasushi Hiraoka
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  9. Hitoshi Kurumizaka
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  10. Wataru Kagawa
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Corresponding authors

Correspondence to Hitoshi Kurumizaka or Wataru Kagawa.

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Yuzurihara, H., Aizawa, Y., Saotome, M. et al. Improved Methods for Preparing the Telomere Tethering Complex Bqt1–Bqt2 for Structural Studies. Protein J 39, 174–181 (2020). https://doi.org/10.1007/s10930-020-09887-z

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  • DOI: https://doi.org/10.1007/s10930-020-09887-z

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