New carbohydrate specificity and HIV-1 fusion blocking activity of the cyanobacterial protein MVL: NMR, ITC and sedimentation equilibrium studies

Carole A. Bewley, Mengli Cai, Satyajit Ray, Rodolfo Ghirlando, Masato Yamaguchi, Koji Muramoto

    研究成果: Article査読

    45 被引用数 (Scopus)

    抄録

    Carbohydrate-binding proteins that bind their carbohydrate ligands with high affinity are rare and therefore of interest because they expand our understanding of carbohydrate specificity and the structural requirements that lead to high-affinity interactions. Here, we use NMR and isothermal titration calorimetry techniques to determine carbohydrate specificity and affinities for a novel cyanobacterial protein, MVL, and show that MVL binds oligomannosides such as Man6GlcNAc2 with sub-micromolar affinities. The amino acid sequence of MVL contains two homologous repeats, each comprising 54 amino acid residues. Using multi-dimensional NMR techniques, we show that MVL contains two novel carbohydrate recognition domains composed of four non-contiguous regions comprising ∼15 amino acid residues each, and that these residues make numerous intermolecular contacts with their carbohydrate ligands. NMR screening of a comprehensive panel of di-, tri-, and high-mannose oligosaccharides establish that high-affinity binding requires at least the presence of a discrete conformation presented by Manβ(1→4)GlcNAc in the context of larger oligomannosides. As shown by sedimentation equilibrium and gel-filtration experiments, MVL is a monodisperse dimer in solution, and NMR data establish that the three-dimensional structure must be symmetric. MVL inhibits HIV-1 Envelope-mediated cell fusion with an IC50 value of ∼30nM.

    本文言語English
    ページ(範囲)901-914
    ページ数14
    ジャーナルJournal of Molecular Biology
    339
    4
    DOI
    出版ステータスPublished - 2004 6 11

    ASJC Scopus subject areas

    • 構造生物学
    • 分子生物学

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