Crystallization of the functional domain of human thrombopoietin using an antigen-binding fragment derived from neutralizing monoclonal antibody

Ryota Kuroki, Masako Hirose, Yoichi Kato, Michael D. Feese, Taro Tamada, Hideki Shigematsu, Hiroshi Watarai, Yoshitake Maeda, Tomoyuki Tahara, Takashi Kato, Hiroshi Miyazaki

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Thrombopoietin (TPO) is a cytokine which primarily stimulates megakaryocytopoiesis and thrombopoiesis. The functional domain of TPO (TPO163) consisting of the N-terminal 163 amino acids was prepared and crystallized. Since the crystallization of TPO163 was unsuccessful using the standard screening methods, a Fab fragment derived from a neutralizing monoclonal antibody was used for crystallization. It was found that the TPO163-Fab complex crystallized reproducibly in 0.1 M potassium phosphate buffer pH 6.0 containing 20-25% polyethylene glycol 4000. Thin crystals (0.2 × 0.2 × 0.02 mm) grew in two space groups: P21, with unit-cell parameters a = 133.20, b = 46.71, c = 191.47 Å, β = 90.24°, and C2, with unit-cell parameters a = 131.71, b = 46.48, c = 184.63 Å, β = 90.42°. The results of a molecular-replacement analysis indicate that the Fab molecules interact with each other and provide a suitable interface for crystallization.

Original languageEnglish
Pages (from-to)856-858
Number of pages3
JournalActa Crystallographica Section D: Biological Crystallography
Volume58
Issue number5
DOIs
Publication statusPublished - 2002
Externally publishedYes

Fingerprint

Thrombopoietin
antigens
Crystallization
Neutralizing Antibodies
antibodies
Thrombopoiesis
Monoclonal Antibodies
fragments
crystallization
Antigens
potassium phosphates
Immunoglobulin Fab Fragments
cells
amino acids
glycols
polyethylenes
Buffers
Screening
screening
buffers

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biophysics
  • Clinical Biochemistry
  • Structural Biology
  • Condensed Matter Physics

Cite this

Crystallization of the functional domain of human thrombopoietin using an antigen-binding fragment derived from neutralizing monoclonal antibody. / Kuroki, Ryota; Hirose, Masako; Kato, Yoichi; Feese, Michael D.; Tamada, Taro; Shigematsu, Hideki; Watarai, Hiroshi; Maeda, Yoshitake; Tahara, Tomoyuki; Kato, Takashi; Miyazaki, Hiroshi.

In: Acta Crystallographica Section D: Biological Crystallography, Vol. 58, No. 5, 2002, p. 856-858.

Research output: Contribution to journalArticle

Kuroki, Ryota ; Hirose, Masako ; Kato, Yoichi ; Feese, Michael D. ; Tamada, Taro ; Shigematsu, Hideki ; Watarai, Hiroshi ; Maeda, Yoshitake ; Tahara, Tomoyuki ; Kato, Takashi ; Miyazaki, Hiroshi. / Crystallization of the functional domain of human thrombopoietin using an antigen-binding fragment derived from neutralizing monoclonal antibody. In: Acta Crystallographica Section D: Biological Crystallography. 2002 ; Vol. 58, No. 5. pp. 856-858.
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AU - Shigematsu, Hideki

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AU - Miyazaki, Hiroshi

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