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Unravelling glucan recognition systems by glycome microarrays using the designer approach and mass spectrometry.

Palma AS, Liu Y, Zhang H, Zhang Y, McCleary BV, Yu G, Huang Q, Guidolin LS, Ciocchini AE, Torosantucci A, Wang D, Carvalho AL, Fontes CM, Mulloy B, Childs RA, Feizi T, Chai W - Mol. Cell Proteomics (2015)

Bottom Line: The glucome microarray comprises 153 oligosaccharide probes with high purity, representing major sequences in glucans.The system is validated using antibodies and carbohydrate-binding modules known to target α- or β-glucans in different biological contexts, extending knowledge on their specificities, and applied to reveal new information on glucan recognition by two signaling molecules of the immune system against pathogens: Dectin-1 and DC-SIGN.The sequencing of the glucan oligosaccharides by the MS method and their interrogation on the microarrays provides detailed information on linkage, sequence and chain length requirements of glucan-recognizing proteins, and are a sensitive means of revealing unsuspected sequences in the polysaccharides.

View Article: PubMed Central - PubMed

Affiliation: From the ‡Glycosciences Laboratory, Department of Medicine, Imperial College London, United Kingdom; §UCIBIO-REQUIMTE, Department of Chemistry, Faculty of Science and Technology, NOVA University of Lisbon; angelina.palma@fct.unl.pt w.chai@imperial.ac.uk.

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Application of the negative-ion ESI-CID-MS/MS method to sequence determination and in detection of unsuspected gluco-oligosaccharide sequences. (A) MS/MS (upper panel) and quasi-MS3 (lower panel) for sequencing a pentasaccharide Barley-5a isolated from barley β-glucan hydrolysate. (B) MS/MS of Bio-Gel P4 fraction of Poria cocos glucan oligosaccharide, Poria-7, before (upper panel) and after (lower panel) removal of minor contaminant by semipreparative HPTLC.
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Figure 4: Application of the negative-ion ESI-CID-MS/MS method to sequence determination and in detection of unsuspected gluco-oligosaccharide sequences. (A) MS/MS (upper panel) and quasi-MS3 (lower panel) for sequencing a pentasaccharide Barley-5a isolated from barley β-glucan hydrolysate. (B) MS/MS of Bio-Gel P4 fraction of Poria cocos glucan oligosaccharide, Poria-7, before (upper panel) and after (lower panel) removal of minor contaminant by semipreparative HPTLC.

Mentions: The pentasaccharide Barley-5a was isolated from barley β-glucan after digestion with a novel cellulase. The sequence was predicted to be Glcβ1,3Glcβ1,4Glcβ1,4Glcβ1,4Glc. However, in addition to 1,4-linkage with ion set -60/78/120 and nonreducing 1,3-linkage with absence of A-fragments, there was an A-type ion set -60/90/120 in spectrum, identifying an internal 1,6-linkage (Fig. 4A). The sequence could be assigned as Glcβ1,3Glcβ1,4Glcβ1,6Glcβ1,4Glc, which was corroborated by NMR (supplemental Results and supplemental Table S4).


Unravelling glucan recognition systems by glycome microarrays using the designer approach and mass spectrometry.

Palma AS, Liu Y, Zhang H, Zhang Y, McCleary BV, Yu G, Huang Q, Guidolin LS, Ciocchini AE, Torosantucci A, Wang D, Carvalho AL, Fontes CM, Mulloy B, Childs RA, Feizi T, Chai W - Mol. Cell Proteomics (2015)

Application of the negative-ion ESI-CID-MS/MS method to sequence determination and in detection of unsuspected gluco-oligosaccharide sequences. (A) MS/MS (upper panel) and quasi-MS3 (lower panel) for sequencing a pentasaccharide Barley-5a isolated from barley β-glucan hydrolysate. (B) MS/MS of Bio-Gel P4 fraction of Poria cocos glucan oligosaccharide, Poria-7, before (upper panel) and after (lower panel) removal of minor contaminant by semipreparative HPTLC.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4390274&req=5

Figure 4: Application of the negative-ion ESI-CID-MS/MS method to sequence determination and in detection of unsuspected gluco-oligosaccharide sequences. (A) MS/MS (upper panel) and quasi-MS3 (lower panel) for sequencing a pentasaccharide Barley-5a isolated from barley β-glucan hydrolysate. (B) MS/MS of Bio-Gel P4 fraction of Poria cocos glucan oligosaccharide, Poria-7, before (upper panel) and after (lower panel) removal of minor contaminant by semipreparative HPTLC.
Mentions: The pentasaccharide Barley-5a was isolated from barley β-glucan after digestion with a novel cellulase. The sequence was predicted to be Glcβ1,3Glcβ1,4Glcβ1,4Glcβ1,4Glc. However, in addition to 1,4-linkage with ion set -60/78/120 and nonreducing 1,3-linkage with absence of A-fragments, there was an A-type ion set -60/90/120 in spectrum, identifying an internal 1,6-linkage (Fig. 4A). The sequence could be assigned as Glcβ1,3Glcβ1,4Glcβ1,6Glcβ1,4Glc, which was corroborated by NMR (supplemental Results and supplemental Table S4).

Bottom Line: The glucome microarray comprises 153 oligosaccharide probes with high purity, representing major sequences in glucans.The system is validated using antibodies and carbohydrate-binding modules known to target α- or β-glucans in different biological contexts, extending knowledge on their specificities, and applied to reveal new information on glucan recognition by two signaling molecules of the immune system against pathogens: Dectin-1 and DC-SIGN.The sequencing of the glucan oligosaccharides by the MS method and their interrogation on the microarrays provides detailed information on linkage, sequence and chain length requirements of glucan-recognizing proteins, and are a sensitive means of revealing unsuspected sequences in the polysaccharides.

View Article: PubMed Central - PubMed

Affiliation: From the ‡Glycosciences Laboratory, Department of Medicine, Imperial College London, United Kingdom; §UCIBIO-REQUIMTE, Department of Chemistry, Faculty of Science and Technology, NOVA University of Lisbon; angelina.palma@fct.unl.pt w.chai@imperial.ac.uk.

Show MeSH
Related in: MedlinePlus