Combinatorial chemoenzymatic synthesis and high-throughput screening of sialosides

Harshal A. Chokhawala, Shengshu Huang, Kam Lau, Hai Yu, Jiansong Cheng, Vireak Thon, Nancy Hurtado-Ziola, Juan A. Guerrero, Ajit Varki, Xi Chen

Research output: Contribution to journalArticlepeer-review

75 Scopus citations

Abstract

Although the vital roles of structures containing sialic acid in biomolecular recognition are well documented, limited information is available on how sialic acid structural modifications, sialyl linkages, and the underlying glycan structures affect the binding or the activity of sialic acid-recognizing proteins and related downstream biological processes. A novel combinatorial chemoenzymatic method has been developed for the highly efficient synthesis of biotinylated sialosides containing different sialic acid structures and different underlying glycans in 96-well plates from biotinylated sialyltransferase acceptors and sialic acid precursors. By transferring the reaction mixtures to NeutrAvidin-coated plates and assaying for the yields of enzymatic reactions using lectins recognizing sialyltransferase acceptors but not the sialylated products, the biotinylated sialoside products can be directly used, without purification, for high-throughput screening to quickly identify the ligand specificity of sialic acid-binding proteins. For a proof-of-principle experiment, 72 biotinylated α2,6-linked sialosides were synthesized in 96-well plates from 4 biotinylated sialyltransferase acceptors and 18 sialic acid precursors using a one-pot three-enzyme system. High-throughput screening assays performed in NeutrAvidin-coated microtiter plates show that whereas Sambucus nigra Lectin binds to α2,6-linked sialosides with high promiscuity, human Siglec-2 (CD22) is highly selective for a number of sialic acid structures and the underlying glycans in its sialoside ligands.

Original languageEnglish (US)
Pages (from-to)567-576
Number of pages10
JournalACS Chemical Biology
Volume3
Issue number9
DOIs
StatePublished - Sep 19 2008

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine

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