Detection and quantification of on-chip phosphorylated peptides by surface plasmon resonance imaging techniques using a phosphate capture molecule

Kazuki Inamori, Motoki Kyo, Yoshiaki Nishiya, Yusuke Inoue, Tatsuhiko Sonoda, Eiji Kinoshita, Tohru Koike, Yoshiki Katayama

研究成果: ジャーナルへの寄稿記事

108 引用 (Scopus)

抄録

We describe herein a detection and quantification system for on-chip phosphorylation of peptides by surface plasmon resonance (SPR) imaging techniques using a newly synthesized phosphate capture molecule (i.e., biotinylated zinc(II) complex). The biotinylated compound is a dinuclear zinc(II) complex that is suitable for accessing phosphate anions as a bridging ligand on the two zinc(II) ions. The compound was exposed on the peptide array and detected with streptavidin (SA) via a biotin-SA interaction by SPR imaging. In the conventional method using antibody, both anti-phosphoserine and anti-phosphotyrosine antibodies were required for phosphoserine and phosphotyrosine detection, respectively. Detection of the phosphate group by the zinc(II) complex, however, was independent of the phosphorylated amino acid residues. The calibration curve for the phosphorylation ratios was established with a calibration chip, on which phosphoserine-containing peptide probes were immobilized. The peptide probes, which were phosphorylated on the surface by protein kinase A, were detected and quantified by SPR imaging using the zinc(II) complex, SA, and anti-SA antibody. The reaction rate and the kinetics of on-chip phosphorylation were also evaluated with the peptide array. The phosphorylation ratio was saturated at ∼20% in 2 h in this study.

元の言語英語
ページ(範囲)3979-3985
ページ数7
ジャーナルAnalytical chemistry
77
発行部数13
DOI
出版物ステータス出版済み - 7 1 2005

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Surface plasmon resonance
Phosphorylation
Streptavidin
Zinc
Phosphoserine
Phosphates
Imaging techniques
Peptides
Molecules
Phosphotyrosine
Calibration
Antibodies
Biotin
Cyclic AMP-Dependent Protein Kinases
Reaction rates
Anions
Anti-Idiotypic Antibodies
Membrane Proteins
Ions
Ligands

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry

これを引用

Detection and quantification of on-chip phosphorylated peptides by surface plasmon resonance imaging techniques using a phosphate capture molecule. / Inamori, Kazuki; Kyo, Motoki; Nishiya, Yoshiaki; Inoue, Yusuke; Sonoda, Tatsuhiko; Kinoshita, Eiji; Koike, Tohru; Katayama, Yoshiki.

:: Analytical chemistry, 巻 77, 番号 13, 01.07.2005, p. 3979-3985.

研究成果: ジャーナルへの寄稿記事

Inamori, Kazuki ; Kyo, Motoki ; Nishiya, Yoshiaki ; Inoue, Yusuke ; Sonoda, Tatsuhiko ; Kinoshita, Eiji ; Koike, Tohru ; Katayama, Yoshiki. / Detection and quantification of on-chip phosphorylated peptides by surface plasmon resonance imaging techniques using a phosphate capture molecule. :: Analytical chemistry. 2005 ; 巻 77, 番号 13. pp. 3979-3985.
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abstract = "We describe herein a detection and quantification system for on-chip phosphorylation of peptides by surface plasmon resonance (SPR) imaging techniques using a newly synthesized phosphate capture molecule (i.e., biotinylated zinc(II) complex). The biotinylated compound is a dinuclear zinc(II) complex that is suitable for accessing phosphate anions as a bridging ligand on the two zinc(II) ions. The compound was exposed on the peptide array and detected with streptavidin (SA) via a biotin-SA interaction by SPR imaging. In the conventional method using antibody, both anti-phosphoserine and anti-phosphotyrosine antibodies were required for phosphoserine and phosphotyrosine detection, respectively. Detection of the phosphate group by the zinc(II) complex, however, was independent of the phosphorylated amino acid residues. The calibration curve for the phosphorylation ratios was established with a calibration chip, on which phosphoserine-containing peptide probes were immobilized. The peptide probes, which were phosphorylated on the surface by protein kinase A, were detected and quantified by SPR imaging using the zinc(II) complex, SA, and anti-SA antibody. The reaction rate and the kinetics of on-chip phosphorylation were also evaluated with the peptide array. The phosphorylation ratio was saturated at ∼20{\%} in 2 h in this study.",
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AU - Sonoda, Tatsuhiko

AU - Kinoshita, Eiji

AU - Koike, Tohru

AU - Katayama, Yoshiki

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