Double-modification of lectin using two distinct chemistries for fluorescent ratiometric sensing and imaging saccharides in test tube or in cell

Eiji Nakata, Yoichiro Koshi, Erina Koga, Yoshiki Katayama, Itaru Hamachi

研究成果: ジャーナルへの寄稿記事

51 引用 (Scopus)

抄録

The site-selective incorporation of two different fluorophores into a naturally occurring protein (lectin, a sugar-binding protein) has been successfully carried out using two distinct orthogonal chemical methods. By post-photoaffinity labeling modification, Con A, a glucose- and mannose-selective lectin, was modified with fluorescein in the proximity of the sugar binding site (Tyr100 site), and the controlled acylation reaction provided the site-selective attachment of coumarin at Lys114. In this doubly modified Con A, the fluorescein emission changed upon the binding to the corresponding sugars, such as the glucose or mannose derivatives, whereas the coumarin emission was constant. Thus, the doubly modified Con A fluorescently sensed the glucose- and mannose-rich saccharides in a ratiometric manner while retaining the natural binding selectivity and affinity, regardless of the double modification. On the benefit of the ratiometric fluorescent analysis using two distinct probes, the sugar trimming process of a glycoprotein can be precisely monitored by the engineered Con A. Furthermore, the doubly modified Con A can be used not only for the convenient fluorescent imaging of saccharides localized on a cell surface, such as the MCF-7, a breast cancer cell having rich high-mannose branch, but also for the ratiometric fluorescent sensing of the glucose concentration inside HepG2 cells. These results demonstrated that the semisynthetic lectin modified doubly by two distinct chemistries is superior to the singly modified one in function, and thus, it may be potentially useful in cell, as well as in test tube.

元の言語英語
ページ(範囲)13253-13261
ページ数9
ジャーナルJournal of the American Chemical Society
127
発行部数38
DOI
出版物ステータス出版済み - 9 28 2005

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Mannose
Lectins
Sugars
Glucose
Imaging techniques
Fluorescein
Coumarins
Acylation
Glycoproteins
Trimming
Fluorophores
Hep G2 Cells
Binding sites
Labeling
Carrier Proteins
Binding Sites
Cells
Breast Neoplasms
Derivatives
Proteins

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

これを引用

Double-modification of lectin using two distinct chemistries for fluorescent ratiometric sensing and imaging saccharides in test tube or in cell. / Nakata, Eiji; Koshi, Yoichiro; Koga, Erina; Katayama, Yoshiki; Hamachi, Itaru.

:: Journal of the American Chemical Society, 巻 127, 番号 38, 28.09.2005, p. 13253-13261.

研究成果: ジャーナルへの寄稿記事

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