Noncovalent microarrays from synthetic amino-terminating glycans: Implications in expanding glycan microarray diversity and platform comparison.

Chunxia Li,Chao Gao,Pengtao Zhang,Lisete M Silva,Angelina S Palma,Zhen Li,Wengang Chai,Peter H Seeberger,Leonid M Likhosherstov,Vladimir Piskarev,Filipa Trovão,Ulrika Westerlind,Markus Weishaupt,Jin Yu,Yibing Zhang

doi:10.1093/glycob/cwab037

Abstract

Glycan microarrays have played important roles in detection and specificity assignment of glycan recognition by proteins. However, the size and diversity of glycan libraries in current microarray systems are small compared to estimated glycomes, and these may lead to missed detection or incomplete assignment. For microarray construction, covalent and noncovalent immobilization are the two types of methods used, but a direct comparison of results from the two platforms is required. Here we develop a chemical strategy to prepare lipid-linked probes from both naturally derived aldehyde-terminating and synthetic amino-terminating glycans that addresses the two aspects: expansion of sequence-defined glycan libraries and comparison of the two platforms. We demonstrate the specific recognition by plant and mammalian lectins, carbohydrate-binding modules and antibodies and the overall similarities from the two platforms. Our results provide new knowledge on unique glycan-binding specificities for the immune receptor Dectin-1 toward β-glucans and the interaction of rotavirus P[19] adhesive protein with mucin O-glycan cores.

Highlights

Recognition of glycans by proteins is crucial to understand molecular mechanisms in health and disease
The Consortium for Functional Glycomics (CFG) and Max Planck Institute (MPI) arrays comprise amino-terminating synthetic glycans that are covalently immobilized on N-hydroxysuccinimide (NHS)-functionalized slides, whereas the NGL arrays comprise mainly naturally derived aldehyde-terminating glycans conjugated to a long chain amino-phospholipid and noncovalently immobilized on nitrocellulose-coated slides
We showed that peanut agglutinin (PNA) lectin, widely used in the detection of core 1 T-antigen, exhibits strong binding to the core 2 antigen and that the O-GlcNAc-specific antibody CTD110.6 and C. mixtus CmCBM6–2 can accommodate core 2 and core 4 antigens, through recognition of the terminal GlcNAcβ1- residues

Summary

Introduction

Recognition of glycans by proteins is crucial to understand molecular mechanisms in health and disease Carbohydrate microarrays, including those of polysaccharides (Wang et al 2002) and sequence-defined glycans (Fukui et al 2002), have played a major role in dissecting glycan–protein interactions after their emergence as a natural follow-up to the development of the microarray technologies for nucleic acids (Schena and Shalon 1995) and proteins (MacBeath and Schreiber 2000). Since their inception in 2002, glycan microarrays have proven to be powerful tools in the detection and specificity assignment of glycan–protein interactions with implications in biology and medicine. The three platforms are in some ways complementary with partial overlap in their glycan repertoires

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