Versatile strategy for the synthesis of biotin-labelled glycans, their immobilization to establish a bioactive surface and interaction studies with a lectin on a biochip

Muñoz, Francisco; Rumbero, Ángel; Sinisterra, J. V.; Santos, J. Ignacio; André, Sabine; Gabius, Hans‐Joachim; Jiménez‐Barbero, Jesús; Hernáiz, María J.

doi:10.1007/s10719-008-9115-y

Cited by 17 publications

(11 citation statements)

References 61 publications

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“…The labelling of sugar chains using 2-aminopyridine (PA) (19,20). As expected, our novel f-mono linker molecule showed fluorescence at an excitation (ex) maximum of 335 nm and an emission (em) maximum of 380 nm.…”

Section: Resultssupporting

confidence: 53%

High-Sensitivity Analysis of Naturally Occurring Sugar Chains, Using a Novel Fluorescent Linker Molecule

Satō

Ito

Arima

et al. 2009

The Journal of Biochemistry

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To analyse the binding of sugar chains to proteins, viruses and cells, the surface plasmon resonance (SPR) technique is very convenient and effective because it is a real-time, non-destructive detection system. Key to this method is linker compounds for immobilization of the sugar chains to the gold-coated chip for SPR. Also, welldesigned fluorescent labelling reagents are essential when analysing the structure of trace amounts of sugar chains derived from natural sources, such as glycoproteins on the surface of specific cells. In this report, we developed a novel linker molecule, named 'f-mono', which has both of these properties: simple immobilization chemistry and a fluorescent label. Since the molecule contains a 2,5-diaminopyridyl group and a thioctic acid group, conjugation with sugar chains can be achieved using the wellestablished reductive amination reaction. This conjugate of sugar chain and fluorescent linker (fluorescent ligand-conjugate, FLC) has fluorescent properties (ex. 335 nm, em. 380 nm), and as little as 1 kg of FLC can be easily purified using HPLC with a fluorescent detector. MS and MS/MS analysis of the FLC is also possible. As a +2 Da larger MS peak ([M + H + 2] + ion) was always associated with the theoretical MS peak ([M + H] + ) (due to the reduction of the thioctic acid moiety), the MS peaks of the FLC were easily found, even using unfractionated crude samples. Immobilization of the FLC onto goldcoated chips, and their subsequent SPR analyses were successively accomplished, as had been performed previously using non-fluorescent ligand conjugates.Key words: immobilization, sugar chain, high sensitivity, analysis, fluorescence, linker molecule, mass spectrometry. The carbohydrates that make up proteoglycans, glycoproteins or glycolipids are responsible for many biological functions and play crucial roles in cellular binding and signalling (1). However, because of their structural complexity, the methods for studying sugar chains are more challenging than that for DNA, RNA or proteins. The numerous isomeric and anomeric configurations of sugar chains, as well as the difficulties in isolating sufficient quantities of naturally occurring sugars, make binding analysis and structure-function studies challenging.For the structural analysis of naturally occurring sugar chains, fluorescent labelling of the sugars has been one popular technique (2). Recently, mass spectrometry (MS) has been used for structural analysis of sugar structures, thanks to the development of structurally well-defined standards (3, 4). Surface plasmon resonance (SPR) methodology is also a very effective method to quantify binding interactions between sugar-chains and lectins or viruses in real time, because it is a non-destructive technology that does not require large quantities of the often scarce materials to be studied (5-9). We have previously reported the development of the 'sugar chip', in which defined sugar chains are immobilized on an SPR sensor chip using our specialized linker molecules (10,11). But the puri...

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Section: Resultssupporting

confidence: 53%

High-Sensitivity Analysis of Naturally Occurring Sugar Chains, Using a Novel Fluorescent Linker Molecule

Satō

Ito

Arima

et al. 2009

The Journal of Biochemistry

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“…Carnosine ethyl ester was synthesized from AH in ethanol at 0°C with acetylchloride. 2-bromoethyl-(2,3,4,6-tetraacetyl)-β-Dglucopyranoside and 2-bromoethyl-(2,3,4,6-tetraacetyl-β-Dgalactopyranosyl)-(1→4)-2,3,6-triacetyl-β-D-glucopyranoside were synthesized as reported elsewhere [41,42].…”

Section: Chemicalsmentioning

confidence: 99%

New glycoside derivatives of carnosine and analogs resistant to carnosinase hydrolysis: Synthesis and characterization of their copper(II) complexes

Lanza

Bellia

D’Agata

et al. 2011

Journal of Inorganic Biochemistry

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“…To avoid the need for fluorescently tagged targets, which are both expensive and demanding to produce, many glycan array methodologies are evaluated using techniques such as Surface Plasmon Resonance (SPR)30, 39–43 and Quartz Crystal Microbalance (QCM)23, 32, 44, 45 which instead rely on the physical properties of surface plasmon absorption and mass change to detect target binding.…”

Section: Introductionmentioning

confidence: 99%

Photo-Click Immobilization on Quartz Crystal Microbalance Sensors for Selective Carbohydrate−Protein Interaction Analyses

et al. 2010

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A photo-click method based on azide photoligation and Cu-catalyzed azide-alkyne cycloaddition has been evaluated for the immobilization of carbohydrates to polymeric materials. The biomolecular recognition properties of the materials have been investigated with regard to applicable polymeric substrates and selectivity of protein binding. The method was used to functionalize a range of polymeric surfaces (polystyrene, polyacrylamide, poly(ethylene glycol), poly(2-ethyl-2-oxazoline) and polypropene) with various carbohydrate structures (based on α-Dmannose, β-D-galactose and N-acetyl-β-D-glucosamine). The functionalized surfaces were evaluated in real-time studies of protein-carbohydrate interactions using a quartz crystal microbalance flow-through system with a series of different carbohydrate-binding proteins (lectins). The method proved to be robust and versatile, resulting in a range of efficient sensors showing high and predictable protein selectivities.

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Versatile strategy for the synthesis of biotin-labelled glycans, their immobilization to establish a bioactive surface and interaction studies with a lectin on a biochip

Cited by 17 publications

References 61 publications

High-Sensitivity Analysis of Naturally Occurring Sugar Chains, Using a Novel Fluorescent Linker Molecule

High-Sensitivity Analysis of Naturally Occurring Sugar Chains, Using a Novel Fluorescent Linker Molecule

New glycoside derivatives of carnosine and analogs resistant to carnosinase hydrolysis: Synthesis and characterization of their copper(II) complexes

Photo-Click Immobilization on Quartz Crystal Microbalance Sensors for Selective Carbohydrate−Protein Interaction Analyses

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