A Neoglycoprotein-Immobilized Fluorescent Magnetic Bead Suspension Multiplex Array for Galectin-Binding Studies

Carbohydrate-protein conjugates have diverse applications. They have been used clinically as vaccines against bacterial infection and have been developed for high-throughput assays to elucidate the ligand specificities of glycan-binding proteins (GBPs) and antibodies. Here, we report an effective pr...

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Bibliographic Details
Published in:Molecules (Basel, Switzerland) Switzerland), 2021-10, Vol.26 (20), p.6194
Main Authors: Zhang, Libo, Yu, Hai, Bai, Yuanyuan, Mishra, Bijoyananda, Yang, Xiaoxiao, Wang, Jing, Yu, Evan B, Li, Riyao, Chen, Xi
Format: Article
Language:English
Subjects:
Antigens
Arrays
Bacterial diseases
Binding
Blood Proteins - chemistry
Blood Proteins - metabolism
Bovine serum albumin
Carbohydrate Conformation
Carbohydrate Sequence
carbohydrate-protein conjugate
Carbohydrates
Conjugates
Enzymes
Fluorescence
Fluorescent Dyes
galectin
Galectin-3
Galectins - chemistry
Galectins - metabolism
Glucose
Glycan
glycan-binding protein
Glycated Serum Albumin
Glycation End Products, Advanced
Glycoproteins
Humans
Immobilization
Immobilized Proteins
lectin
Lectins
Magnetic Phenomena
Magnetic properties
Mammals
multiplex assay
Multiplexing
Plant Lectins - chemistry
Plant Lectins - metabolism
Polysaccharides - chemistry
Protein Array Analysis - methods
Protein Binding
Proteins
Recombinant Proteins - chemistry
Recombinant Proteins - metabolism
Serum Albumin
Serum Albumin, Bovine
Streptococcus infections
Vaccines
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Summary:Carbohydrate-protein conjugates have diverse applications. They have been used clinically as vaccines against bacterial infection and have been developed for high-throughput assays to elucidate the ligand specificities of glycan-binding proteins (GBPs) and antibodies. Here, we report an effective process that combines highly efficient chemoenzymatic synthesis of carbohydrates, production of carbohydrate-bovine serum albumin (glycan-BSA) conjugates using a squarate linker, and convenient immobilization of the resulting neoglycoproteins on carboxylate-coated fluorescent magnetic beads for the development of a suspension multiplex array platform. A glycan-BSA-bead array containing BSA and 50 glycan-BSA conjugates with tuned glycan valency was generated. The binding profiles of six plant lectins with binding preference towards Gal and/or GalNAc, as well as human galectin-3 and galectin-8, were readily obtained. Our results provide useful information to understand the multivalent glycan-binding properties of human galectins. The neoglycoprotein-immobilized fluorescent magnetic bead suspension multiplex array is a robust and flexible platform for rapid analysis of glycan and GBP interactions and will find broad applications.
ISSN:1420-3049
1420-3049
DOI:10.3390/molecules26206194