Glycomics: 416 (Methods in Enzymology)
Recent avian H5N1 viruses exhibit increased propensity for acquiring human receptor specificity. Galectins-1, -2 and -3 exhibit differential recognition of sialylated glycans and blood group antigens. Dimeric galectin-8 induces phosphatidylserine exposure in leukocytes through polylactosamine recognition by the carboxyl terminal domain. Functional characterization of HFR-1, a high mannose N-glycan-specific wheat lectin induced by Hessian fly larvae. Lectin microarrays identify cell-specific and functionally significant cell surface glycan markers.
Role of cellular heparan sulfate proteoglycans in infection of human adenovirus serotype 3 and Solution structure and sugar-binding mechanism of mouse latrophilin-1 RBL: Dependence of surface monoclonal antibody binding on dynamic changes in FcgammaRIIb expression. Replication and transmission of H9N2 influenza viruses in ferrets: Temporal changes in the carbohydrates expressed on BG01 human embryonic stem cells during differentiation as embryoid bodies.
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Biocompatible carbon nanotubes generated by functionalization with glycodendrimers. The crystal structure of avian CD1 reveals a smaller, more primordial antigen-binding pocket compared to mammalian CD1. Crystal structures of mouse CD1d-iGb3 complex and its cognate Valpha14 T cell receptor suggest a model for dual recognition of foreign and self glycolipids. Siglec-F antibody administration to mice selectively reduces blood and tissue eosinophils.
Glycan microarray analysis of Candida glabrata adhesin ligand specificity. Crystal structures of the staphylococcal toxin SSL5 in complex with sialyl Lewis X reveal a conserved binding site that shares common features with viral and bacterial sialic acid binding proteins. Mechano-transduction mediated secretion and uptake of galectin-3 in breast carcinoma cells: Implications in the extracellular functions of the lectin. Dendritic cell maturation results in pronounced changes in glycan expression affecting recognition by siglecs and galectins. Affinity of galectin-8 and its carbohydrate recognition domains for ligands in solution and at the cell surface.
The crystal structure of staphylococcal superantigen-like protein 11 in complex with sialyl Lewis X reveals the mechanism for cell binding and immune inhibition. Glycolipid and ganglioside metabolism imbalances in Huntington's disease. Phage escape libraries for checkmate analysis. Mechanism of botulinum neurotoxin B and G entry into hippocampal neurons. Characteristics of two CDrelated cell-surface expressed antigens of human lymphocytes. Scavenger receptor C-type lectin binds to the leukocyte cell surface glycan Lewis x by a novel mechanism. The structure of human 4F2hc ectodomain provides a model for homodimerization and electrostatic interaction with plasma membrane.
Effective replication of human influenza viruses in mice lacking a major alpha2,6 sialyltransferase. Clustered carbohydrates as a target for natural killer cells: Receptor binding specificity of recent human H3N2 influenza viruses. Critical functions of N-glycans in L-selectin-mediated lymphocyte homing and recruitment. Clathrate nanostructures for mass spectrometry. Normal human serum contains high levels of anti-GalalphaGlcNAc antibodies.
The liverwort Marchantia polymorphia expresses orthologs of the fungal agaricus bisporus agglutinin family.
Application of Microarrays for Deciphering the Structure and Function of the Human Glycome
A ratiometric lectin microarray approach to analysis of the dynamic mammalian glycome. Expert Opin Biol Ther. The threonine that carries fucose, but not fucose, is required for Cripto to facilitate Nodal signaling. An improved scoring scheme for predicting glycan structures from gene expressison data. Distinct endocytic mechanisms of CD22 Siglec-2 and Siglec-F reflect roles in cell-signaling and innate immunity.
Sialic acid on herpes simplex virus type-1S envelope glycoproteins is required for effeicient infection of Cells. Using computer applications and online resources to teach and learn pharmaceutical chemistry. An Example of Neofunctionalization in Legumes. Phylogenetic and specificity studies of two-domain GNA-related lectins: Evidence for carbohydrate recognition and homotypic and heterotypic binding by the TIM family.
Immunization by avian h5 influenza hemagglutinin mutants with altered receptor binding specificity.
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Siglec-7 undergoes a major conformational change when complexed with the a 2,8-disialylganglioside GT1b. Probing the cis interactions of the inhibitory receptor Siglec-7 with a 2,8-disialylated ligands on natural killer cells and other leukocytes using glycan-specific antibodies and by analysis of a 2,8-sialyltransferase gene expression.
Role of lipid trimming and CD1 groove size in cellular antigen presentation. Galectin-3 associates with the primary cilium and modulates cyst growth in congenital polycystic kidney disease. High-affinity ligand probes of CD22 overcome the threshold set by cis ligands to allow for binding, endocytosis, and killing of B cells. A focused microarray approach to functional glycomics: Two categories of mammalian galactose-binding receptors distinguished by glycan array profiling.
N-terminal truncation enables crystallization of the receptor-binding domain of the FedF bacterial adhesin. Selective deficits in the expression of striatal-enriched mRNAs in Huntingtons disease. Detection of differentially expressed glycogenes in trabecular meshwork of eyes with primary open-angle glaucoma. Evolution of the receptor binding phenotype of influenza A H5 viruses. Regulation of B cell development and B cell signaling by CD22 and its ligands a 2,6-linked sialic acids. Automatic determination of O-glycan structure from fragmentation spectra.
One-step biotinylation procedure for carbohydrates to study carbohydrate-protein interactions. Detection and purification of tyrosine-sulfated proteins using a novel anti-sulfotyrosine monoclonal antibody. Lectin from the Manila clam Ruditapes philippinarum is induced upon infection with the protozoan parasite Perkinsus olseni. Natural killer T cells recognize diacylglycerol antigens from pathogenic bacteria. A gram distribution kernel applied to glycan classification and motif extraction. Role of the carbohydrate recognition domains of mouse galectin-4 in oligosaccharide binding and epitope recognition and expression of galectin-4 and galectin-6 in mouse cells and tissues.
Int J Mol Med. The carbohydrate recognition domain of Dectin-2 is a C-type lectin with specificity for high-mannose. Letter to the Glyco-Forum: Identification of the sialic acid structures recognized by minute virus of mice and the role of binding affinity in virulence adaptation. Altered granulopoietic profile and exaggerated acute neutrophilic inflammation in mice with targeted deficiency in the sialyltransferase, ST6Gal Expression of aberrant forms of CD22 on B lymphocytes in Cd22a lupus-prone mice affects ligand binding.
Search for additional influenza virus to cell interactions. Targeting glycosylation pathways and the cell cycle: Biochemical and structural analysis of Helix pomatia agglutinin HPA: A hexameric lectin with a novel fold. Carbohydrate specificity of an insecticidal lectin isolated from the leaves of Glechoma hederacea towards mammalian glycoconjugates. Genomic and metabolic studies of the impact of probiotics on a model gut symbiont and host. Glycan microarray analysis of the hemagglutinins from modern and pandemic influenza viruses reveals different receptor specificities.
Structure and receptor specificity of the hemagglutinin from an H5N1 influenza virus. Galectin-3 and Galectin-1 bind distinct cell surface glycoprotein receptors to induce T cell death. Large-scale chemoenzymatic synthesis of blood group and tumor-associated poly- N -acetyllactosamine antigens. Design of natural killer T cell activators: Structure and function of a microbial glycosphingolipid bound to mouse CD1d.
Alpha2,3 and alpha2,6 N-linked sialic acids facilitate efficient binding and transduction by adeno-associated virus types 1 and 6. B cell receptors in TCL1 transgenic mice resemble those of aggressive, treatment-resistant human chronic lymphocytic leukemia. Structural characterization of mycobacterial phosphatidylinositol mannoside binding to mouse CD1d. Glycan array screening reveals a candidate ligand for siglec Impact of natural variation in bacterial F17G adhesins on crystallization behaviour.
Acta Crystallogr D Biol Crystallogr. Suppression of tumor formation in lymph nodes by L-selectin-mediated natural killer cell recruitment. Selective binding of the scavenger receptor C-type lectin to Lewis x trisaccharide and related glycan ligands. Synthesis of a phenyl thio-beta-D-galactopyranoside library from 1,5-difluoro-2,4-dinitrobenzene: Receptor-binding properties of swine influenza viruses isolated and propagated in MDCK cells.
Receptor specificity of influenza viruses from birds and mammals: Crystal structure of mouse CD1d bound to the self ligand phosphatidylcholine: Beta1,3-N-acetylglucosaminyltransferase 1 glycosylation is required for axon pathfinding by olfactory sensory neurons. Prediction of glycan structures from gene expression data based on glycosyltransferase reactions.
N-acetylglucosamineO-sulfotransferases 1 and 2 cooperatively control lymphocyte homing through L-selectin ligand biosynthesis in high endothelial venules. Recognition of bacterial glycosphingolipids by natural killer T cells. Dimeric galectin-1 binds with high affinity to alpha2,3-sialylated and non-sialylated terminal N-acetyllactosamine units on surface-bound extended glycans. Synthetic studies of pseurotin A: Fluorescent assay for studying the substrate specificity of neuraminidase. Dietary and genetic control of glucose transporter 2 glycosylation promotes insulin secretion in suppressing diabetes.
Efficient recruitment of lymphocytes in inflamed brain venules requires expression of cutaneous lymphocyte antigen and fucosyltransferase-VII. Schwartz-Albiez R, Kniep B. Perspectives for establishment and reorganization of carbohydrate-directed CD antibodies. Report of the carbohydrate section. Gene expression profiling of mouse postnatal cerebellar development using oligonucleotide microarrays designed to detect differences in glycoconjugate expression.
Mouse Siglec-F and human Siglec-8 are functionally convergent paralogs that are selectively expressed on eosinophils and recognize 6-sulfo-sialyl Lewis X as a preferred glycan ligand. Carbohydrate profiling reveals a distinctive role for the C-type lectin MGL in the recognition of helminth parasites and tumor antigens by dendritic cells. Synthesis of deoxy and acylamino derivatives of lactose and use of these for probing the active site of Neisseria meningitidis N-acetylglucosaminyltransferase.
Versatile fluorescent derivatization of glycans for glycomic analysis. Structure and function of a potent agonist for the semi-invariant natural killer T cell receptor. Molecular mechanism of lipopeptide presentation by CD1a. Structural basis for CD1d presentation of a sulfatide derived from myelin and its implications for autoimmunity.
Sialyltransferase ST8Sia-II assembles a subset of polysialic acid that directs hippocampal axonal targeting and promotes fear behavior. Printed covalent glycan array for ligand profiling of diverse glycan binding proteins. Nat Struct Mol Biol. T Cell Activation by Lipopeptide Antigens. The T cell antigen receptor expressed by V a 14 i NKT cells has a unique mode of glycosphingolipid antigen recognition.
Human galectin-1 recognition of poly-N-acetyllactosamine and chimeric polysaccharides. The first direct evaluation of the two-active site mechanism for chitin synthase. Second-generation dimeric inhibitors of chitin synthase. Expression patterns of alpha 2,3-sialyltransferases and alpha 1,3-fucosyltransferases determine the mode of sialyl Lewis X inhibition by disaccharide decoys. Probing the mechanism of a fungal glycosyltransferase essential for cell wall biosynthesis. UDP-chitobiose is not a substrate for chitin synthase. Transitional and marginal zone B cells have a high proportion of unmasked CD A potent and highly selective inhibitor of human a -1,3-fucosyltransferase via click chemistry.
Crystal structure of CD1a in complex with a sulfatide self antigen at a resolution of 2. Analysis of result variability from high-density oligonucleotide arrays comparing same-species and cross-species hybridizations. Conditional control of selectin ligand expression and global fucosylation events in mice with a targeted mutation at the FX locus.
Glycosylation of mouse and human immune cells: Narayan S, Thomas EA. Sphingolipid abnormalities in psychiatric disorders: Production and application of glycan microarrays. Glycomics hits the big time. Glycan-based high-affinity ligands for toxins and pathogen receptors. Li J, Richards JC. Functional glycomics and glycobiology: Carbohydrate moieties as vaccine candidates: Transcript analysis of stem cells. Mass spectrometric analysis of mutant mice. Multivalent ligands for siglecs. Engineered carbohydrate-recognition domains for glycoproteomic analysis of cell surface glycosylation and ligands for glycan-binding receptors.
Use of glycan microarrays to explore specificity of glycan-binding proteins. Glycans as receptors for influenza pathogenesis. Endogenous galectins and the control of the host inflammatory response. Mass spectrometry in the analysis of N-linked and O-linked glycans. Curr Opin Struct Biol. Turning 'sweet' on immunity: J Comput Sci Syst Biol.
Schiefner A, Wilson IA. Presentation of lipid antigens by CD1 glycoproteins. Context-specific target definition in influenza a virus hemagglutinin-glycan receptor interactions. Symbol nomenclature for glycan representation. New development of glycan arrays. An introduction to bioinformatics for glycomics research.
Characterizing the glycome of the mammalian immune system. Curr Opin Chem Biol. A unified vision of the building blocks of life. Mammalian glycosylation in immunity. Protein-glycan interactions in the control of innate and adaptive immune responses. The chloroform was then removed under a stream of nitrogen. Partially methylated alditol acetates were prepared from permethylated samples for GC-MS linkage analysis as described previously The potential difference between the source acceleration voltage and the collision cell was set to 1 kV, and argon was used as collision gas.
The MALDI was tuned to specifically allow the transmission of higher mass ions at the expense of those at lower mass by manipulation of the lens voltages within Source 1. To prepare samples for mass spectrometry, washed cell pellets from wild type CHO cells grown in monolayer or suspension, as well as nine lectin-resistant mutants Lec1, Lec2, Lec3. Preparation of tryptic glycopeptides facilitates the release of N - and O -glycans by means of peptide N -glycosidase F digestion and reductive elimination, respectively.
Permethylation of the purified glycan populations was employed to enhance the sensitivity of the mass spectrometric analysis and to direct the fragmentation within tandem MS analyses. Although not fully quantitative, recent studies have succeeded in demonstrating that MALDI-MS analyses of permethylated glycans provides reliable relative quantitation information based on signal intensities, particularly when comparisons are made over a small mass range of single ion peaks in the same spectrum Linkage analysis by GC-MS, in concert with the mass spectrometric profiling information, facilitates the assignment of primary structural aspects, such as glycosidic bond positions and monosaccharide constituents.
The assignments of N -glycan spectra were carried out with the assistance of Cartoonist 11 , 15 , a bespoke algorithm designed to mimic the human approach to the analysis and assignment of N -glycan MALDI spectra. Cartoonist searches the raw MS data for peak envelopes and uses knowledge of the biosynthetic pathways to present the user with the most likely permethylated carbohydrate structures for each signal.
The graphical interface of GlycoWorkbench provides an environment in which structure models can be rapidly assembled, automatically matched with MS n data, and compared to assess the best candidate. Due to the finding that CHO cells make not only multiantennary structures but also highly extended LacNAc repeats, the annotations are simplified throughout by using biantennary structures with the extensions listed in parentheses.
Unless specifically noted, such structures are present as a mixture of bi-, tri-, and tetra-antennary glycans with varying numbers of LacNAc extensions and terminal NeuAc, NeuGc, or Fuc. When discussed in the text, the total number of LacNAc units added to the pentasaccharide chitobiose core is given. A mass spectrum obtained from the peptide N -glycosidase F-released and -permethylated N -glycans from a typical monolayer preparation of cells is shown in Fig. Complete compositional assignments can be found in supplemental Tables S1 and S2.
For complete annotation of the spectrum, see supplemental Table S1. Very little evidence was observed for the presence of hybrid N -glycans in wild type CHO samples.
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Sialic acid was predominantly in the form of NeuAc, although NeuGc was present in a small proportion of sialylated N -glycans. It is advantageous that most CHO sialic acid is NeuAc because NeuGc is not desirable in recombinant therapeutic glycoproteins due to the high levels of circulating anti-NeuGc antibodies in human serum By contrast, there was no evidence of N -glycans with more than one fucose residue.
Application of Microarrays for Deciphering the Structure and Function of the Human Glycome
The main difference between the monolayer and a suspension of CHO preparations was a variation in the level of sialylation, with the sCHO preparation showing a slight decrease in relative abundance of sialylated N -glycans compared with the mCHO sample. These peaks are highlighted in green in Fig.
As was the case for the N -glycans, both the mCHO and sCHO O -glycan spectra were very similar, although the cells grown in monolayer again have a slightly higher degree of relative sialylation. Overall, however, the mass at Assignments of the fragment ions generated and alternative antennal arrangements are shown.
The ion highlighted in red represents the cleavage of multiple antennae in a fashion that does not facilitate clear annotation. The presence of these abundant signals, along with the lack of any 3,6-linked galactose in linkage analysis data, confirms that polyLacNAc chains were linear, rather than branched. Additionally, information regarding the number of antennae that are extended by polyLacNAc chains can be inferred by using signals corresponding to the enzyme-trimmed core structures.
Many N -glycans carry a core fucose, and none carry more than one fucose. CHO cell mutants that belong to the Lec2 complementation group have a mutation in the Slc35a1 open reading frame and are unable to translocate CMP-sialic acid into the lumen of the Golgi apparatus, resulting in markedly reduced levels of sialylation of glycoproteins and gangliosides The N -glycans from Lec2 cells were consistent with the mutation, and no sialylated species were observed supplemental Table S1.
This is interesting because previous analyses have shown that glycoconjugates made by Lec2 cells have a small amount of sialic acid 24 , The O -glycan profile of the Lec2 CHO mutant data not shown was similarly affected by the alteration in sialylation of glycoconjugates. For complete annotation of the spectra, see supplemental Table S1.
Lec13 mutant CHO cells exhibit reduced expression of the Gmd gene, resulting in a decrease in GDP-Man-4,6-dehydratase activity and concomitant decreases in levels of GDP-fucose and fucosylated glycans 26 , — , There is a dramatic decrease in the level of fucosylated N -glycans in Lec13, with an increase in the relative abundance of the related, nonfucosylated N -glycans. The wild type sCHO sample shows an approximate fucosylated-to-nonfucosylated ratio of 5.
Nevertheless, fucose was transferred to Lec13 N -glycans Fig. Consistent with this, Lec13 cells bound a small amount of the lectins Pisum sativum agglutinin and Aleuria aurantica lectin, even when grown in dialyzed serum supplemental Fig. However, Lec13 cells did not bind antibodies that recognize fucose on N -glycan antennae supplemental Fig. Other studies have reported small amounts of GDP-fucose in Lec13 cells The ratios of the abundance of N -glycans carrying multiple LacNAc units and thus probable multiantennary N -glycans were characteristically different, with a shift toward potential bi- and tri-antennary N -glycans rather than putative tetra-antennary N -glycans.
These peaks are highlighted in green to aid comparison in Fig. The ratio has shifted from 3: Glycan determinants defined by glycan-binding proteins.
The determinants defined by six commercially available lectins and five monoclonal antibodies were determined by analysis on the defined glycan microarray provided by the Consortium for Functional Glycomics. The determinants, whose presence on an array can be defined by positive signals in an array analysis, are outlined in each structure.
Con A , concanavalin A. The binding patterns of the different lectins can provide extensive structural information and can be extremely useful for differentiating glycans that have the same mass but a different arrangement of monosaccharides. Interestingly, no two patterns were identical despite the fact that all of the glycans were biantennary N -glycans with the same composition. Ten hypothetical N -glycans 1—10 are listed, and the predicted binding patterns for the hypothetical microarray of the lectins and anti-glycan antibodies described in Fig. For example, glycans 6 and 7 Fig.
The positive MAL binding together with antibody binding confirmed the branched structure because these reagents are specific for terminal structures. The data, however, cannot determine on which branch each determinant resides. Space does not permit the interpretation of each data point, but a large amount of data can be associated with each glycan on this example array. These metadata can be compiled in a database and used for predicting monosaccharide sequence and detailed structures of each individual glycan. We have termed this approach metadata-assisted glycan sequencing MAGS.
As the SGM is interrogated with defined GBPs as well as GBPs whose specificity and function are unknown, a database continues to be populated with information on each glycan. When a glycan is determined to be biologically relevant based on a binding event, additional information may be obtained by retrieving the glycan from the TGL for further analysis, although structural information on the entire glycome on the SGM can be addressed by evaluating the binding profile of defined GBPs before and after specific in situ exoglycosidase digestion on the arrays Metadata-assisted glycan sequencing is an extension of the shotgun glycan microarray concept.
Beginning with the generation of the TGL, each glycan is assigned an accession number and printed on the array, and the metadata are collected for each glycan and stored in a database. Pre-printing information can include the following: A variety of techniques has been applied to solve the structures of the complex mixture of isomeric glycans found in human milk We elected to take a functional approach to the human milk-soluble glycome using shotgun glycomics The neutral, monosialyl, and disialyl glycans of a milk sample were tagged with AEAB, separated by two-dimensional HPLC into nearly homogeneous but not fully characterized glycans that made up the human milk TGL.
To demonstrate the utility of the human milk glycan HMG array, we interrogated it with a variety of well characterized GBPs, including lectins and specific anti-glycan antibodies. The other six lectins, A. In interrogations to identify the function of HMGs, we discovered a number of interesting features of these glycans. Some glycans contain epitopes for the monoclonal antibodies TRA-1—60 and TRA-1—81, which are specific for biomarkers of human embryonic pluripotent stem cells Other specifically sialylated glycans are bound by fluorescently labeled influenza A virus and minute virus of mice MVM , suggesting that HMGs may function as receptor decoys in an innate defense mechanism against potential pathogens Whereas molecular mass data provided compositions of the natural glycan ligands bound by these potential pathogens, more detailed MS n data were unable to definitively solve the structures.
To obtain more decisive structural characterizations and demonstrate the utility of the MAGS, we retrieved 22 functionally identified glycans from the TGL of the HM-SGM and printed them with 17 defined milk glycan standards. Individual arrays were interrogated with eight lectins and five anti-glycan monoclonal antibodies that had been analyzed on the CFG glycan microarray to confirm their specificity and binding activity.
Such an analysis is essential in these studies due to the lack of vendor quality control of these reagents. After obtaining the initial patterns of binding, the subarrays were subjected to digestion with specific exoglycosidases either independently or in combination and subsequently interrogated again with the appropriate lectins or anti-glycan antibodies with positive and negative binding indicating the presence or absence of the corresponding determinants.
The results of these studies are summarized in heat maps along with detailed descriptions of the logic behind the assignments of predicted structures of the functionally identified glycans Because of space limitations, the detailed analysis of a single disialylated glycan identified as a glycan ligand bound by MVM is provided here as an example of a structural determination using the MAGS approach.
However, the structural data of 22 HMGs on the array were obtained simultaneously. Several assumptions can be made regarding HMGs based on previous studies 93 , 95 , The results of interrogation of the disialylated glycan and 10 standard milk glycans with a selection of defined lectins and antibodies are shown in Fig. The unknown glycan predicted structure shown in Fig. ECL binding was weak data not shown and was presumably due to the steric effect of the sialylated branch, because the ECL binding signal increased by 3-fold after neuraminidase treatment data not shown , and one branch must be a type 1 structure, which is not bound by ECL.
In addition, anti-type 1 antibody binding is observed only after removal of all the sialic acid by nonspecific neuraminidase. Taken together, the MAGS analysis permits a relatively conclusive prediction that the isolated glycan ligand bound by MVM is as shown in the unknown structure in Fig. Example of MAGS of a single human milk glycan selected for structural analysis based on its binding function. A , lectin and antibody binding to standards and an unknown disialyl human milk glycan.
The predicted structure of the unknown glycan identified as a glycan ligand for MVM from human milk is shown at the upper left above a list of 10 glycan standards obtained from human milk. B , lectin and antibody binding to standards and an unknown disialyl human milk glycan before and after exoglycosidase digestion.
Using this conceptual approach, we were able to propose structures, including most of the linkage positions and anomeric configurations for the glycans that bound TRA-1 antibodies, influenza A, and MVM Some redundancy of structures was observed due to the overlap of glycan fractions collected during multidimensional chromatography. Nevertheless, the correlation of proposed structures with the function defined by antibody or virus binding provided information on the glycans in human milk that are related to embryonic stem cell-specific epitopes and potential receptors for viruses.
Specific lectins are commonly used for the immunohistochemical localization of specific glycan structures in cells and tissues, and with MAGS we simply apply this approach to libraries of structurally undefined glycans with the ability to obtain additional material from the reserved TGL entries for deeper structural characterization. Although the MAGS approach to determining glycan structure alone cannot unequivocally identify a glycan, it can certainly complement MS in glycan structural determination, because it incorporates additional chemical and biochemical approaches used to define glycan structure, including digestions with specific exo-glycosidases, and specific lectin and antibody interactions.
The concept of collecting metadata on structural analysis arrays is rational, and it is not unreasonable to consider the possibility that this approach with miniaturization, increased detection sensitivity, and automation could be developed as a high throughput approach to glycomics analysis. Although MAGS represents a novel approach to assist in high throughput sequencing of glycans, it is highly dependent upon the availability of well characterized GBPs, such as antibodies and lectins, of defined specificity.
Although many such GBPs are available, the quality control of commercial reagents is sometimes lacking; thus, researchers unaware of this may draw incorrect conclusions without independent verification of the activity and specificity of the commercial GBP.