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Applied Biochemistry and Biotechnology

, Volume 68, Issue 1–2, pp 41–56 | Cite as

Synthesis of N-acetylneuraminyl-α2,3(6)lactose-malate dehydrogenase conjugate for detecting sialic acid terminal groups on glycoproteins via homogeneous lectin-based enzyme-linked binding assay

  • Xuan Guo
  • Mark E. Meyerhoff
Original Articles

Abstract

AnN-acetylneuraminyl-α2,3(6)lactose-malate dehydrogenase (MDH-Lac-Neu5Ac) conjugate is prepared via an isothiocyanate conjugation method using ap-aminophenethylamino derivative of sialyllactose. The newly synthesized conjugate can be utilized as a reagent in a novel homogeneous lectin-based, enzyme-linked, competitive binding assay (1–3) for probing the specific carbohydrate structure and content of intact glycoproteins. The enzymatic activity of the MDH-Lac-Neu5Ac conjugate is shown to be significantly inhibited (35%) by sialic acid-binding lectin,Limax flavus agglutinin (LFA), and this inhibition is reversed by mucin, a glycoprotein possessing sialic acid terminals. The asialo form of mucin, however, binds weakly to LFA, yielding no substantial increase in the MDH-Lac-Neu5Ac activity at comparable glycoprotein concentrations. Use of the newly synthesized conjugate in conjunction with LFA or other lectins capable of binding sialic acid may provide a rapid and convenient way to detect the presence and relative amount of sialic acid terminal groups within intact glycoprotein structures.

Index Entries

Sialic acid malate dehydrogenase conjugate homogeneous binding assay glycoproteins 

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Copyright information

© Humana Press Inc 1997

Authors and Affiliations

  • Xuan Guo
    • 1
  • Mark E. Meyerhoff
    • 1
  1. 1.Department of ChemistryUniversity of MichiganAnn Arbor

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