Characterisation of the wheat germ agglutinin binding to self-assembled monolayers of neoglycoconjugates by AFM and SPR

Michael Lienemann, Arja Paananen, Harry Boer, Jesús M. de la Fuente, Isabel García, Soledad Penadés, Anu Koivula

Research output: Contribution to journalArticleScientificpeer-review

19 Citations (Scopus)

Abstract

Carbohydrate–protein interactions govern many crucial life processes involved in cell recognition events, but are often difficult to study because the interactions are weak, and multivalent exposure appears to be crucial for their biological function. We have used self-assembled monolayers (SAMs) of neoglycoconjugates as a model system to probe the specific interactions between the lectin wheat germ agglutinin (WGA) and monosaccharides by surface plasmon resonance (SPR) and atomic force microscopy (AFM) force measurements. SAMs presenting N-acetyl-D-glucosamine (GlcNAc) as a neoglycoconjugate were produced on gold surfaces, where the SAM formation was monitored using a quartz crystal microbalance (QCM) and shown to be a very rapid process. In the AFM force measurements WGA was covalently coupled to flexible polyethylene glycol (PEG) molecules at a probe surface using amine coupling. GlcNAc-specific binding events were detected with a WGA-modified probe on the GlcNAc-neoglycoconjugate SAM at bond rupture forces of 47 ± 15 pN. Additionally, less frequent GlcNAc-specific unbinding events were detected at higher forces (120 ± 20 pN) which are believed to originate from simultaneous detachment of multiple binding sites from the SAM surface. SPR measurements confirmed that WGA has higher affinity toward the immobilized GlcNAc-SAM than toward the soluble free monosaccharide. The binding constants obtained for soluble chitinoligosaccharides suggested up to three subsites within one carbohydrate-binding site of the WGA molecule and also provided further evidence of the multivalent binding character of the WGA dimer.
Original languageEnglish
Pages (from-to)633 - 643
JournalGlycobiology
Volume19
Issue number6
DOIs
Publication statusPublished - 24 Feb 2009
MoE publication typeA1 Journal article-refereed

Fingerprint

Wheat Germ Agglutinins
Surface Plasmon Resonance
Atomic Force Microscopy
Surface plasmon resonance
Self assembled monolayers
Atomic force microscopy
Monosaccharides
Force measurement
Binding Sites
Quartz Crystal Microbalance Techniques
Molecules
Acetylglucosamine
Quartz crystal microbalances
Lectins
Gold
Dimers
Amines
Rupture
Carbohydrates

Keywords

  • AFM force spectroscopy
  • protein-carbohydrate interaction
  • self-assembled monolayer
  • surface plasmon resonance
  • wheat germ agglutinin

Cite this

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title = "Characterisation of the wheat germ agglutinin binding to self-assembled monolayers of neoglycoconjugates by AFM and SPR",
abstract = "Carbohydrate–protein interactions govern many crucial life processes involved in cell recognition events, but are often difficult to study because the interactions are weak, and multivalent exposure appears to be crucial for their biological function. We have used self-assembled monolayers (SAMs) of neoglycoconjugates as a model system to probe the specific interactions between the lectin wheat germ agglutinin (WGA) and monosaccharides by surface plasmon resonance (SPR) and atomic force microscopy (AFM) force measurements. SAMs presenting N-acetyl-D-glucosamine (GlcNAc) as a neoglycoconjugate were produced on gold surfaces, where the SAM formation was monitored using a quartz crystal microbalance (QCM) and shown to be a very rapid process. In the AFM force measurements WGA was covalently coupled to flexible polyethylene glycol (PEG) molecules at a probe surface using amine coupling. GlcNAc-specific binding events were detected with a WGA-modified probe on the GlcNAc-neoglycoconjugate SAM at bond rupture forces of 47 ± 15 pN. Additionally, less frequent GlcNAc-specific unbinding events were detected at higher forces (120 ± 20 pN) which are believed to originate from simultaneous detachment of multiple binding sites from the SAM surface. SPR measurements confirmed that WGA has higher affinity toward the immobilized GlcNAc-SAM than toward the soluble free monosaccharide. The binding constants obtained for soluble chitinoligosaccharides suggested up to three subsites within one carbohydrate-binding site of the WGA molecule and also provided further evidence of the multivalent binding character of the WGA dimer.",
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author = "Michael Lienemann and Arja Paananen and Harry Boer and {de la Fuente}, {Jes{\'u}s M.} and Isabel Garc{\'i}a and Soledad Penad{\'e}s and Anu Koivula",
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Characterisation of the wheat germ agglutinin binding to self-assembled monolayers of neoglycoconjugates by AFM and SPR. / Lienemann, Michael; Paananen, Arja; Boer, Harry; de la Fuente, Jesús M.; García, Isabel; Penadés, Soledad; Koivula, Anu.

In: Glycobiology, Vol. 19, No. 6, 24.02.2009, p. 633 - 643.

Research output: Contribution to journalArticleScientificpeer-review

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T1 - Characterisation of the wheat germ agglutinin binding to self-assembled monolayers of neoglycoconjugates by AFM and SPR

AU - Lienemann, Michael

AU - Paananen, Arja

AU - Boer, Harry

AU - de la Fuente, Jesús M.

AU - García, Isabel

AU - Penadés, Soledad

AU - Koivula, Anu

PY - 2009/2/24

Y1 - 2009/2/24

N2 - Carbohydrate–protein interactions govern many crucial life processes involved in cell recognition events, but are often difficult to study because the interactions are weak, and multivalent exposure appears to be crucial for their biological function. We have used self-assembled monolayers (SAMs) of neoglycoconjugates as a model system to probe the specific interactions between the lectin wheat germ agglutinin (WGA) and monosaccharides by surface plasmon resonance (SPR) and atomic force microscopy (AFM) force measurements. SAMs presenting N-acetyl-D-glucosamine (GlcNAc) as a neoglycoconjugate were produced on gold surfaces, where the SAM formation was monitored using a quartz crystal microbalance (QCM) and shown to be a very rapid process. In the AFM force measurements WGA was covalently coupled to flexible polyethylene glycol (PEG) molecules at a probe surface using amine coupling. GlcNAc-specific binding events were detected with a WGA-modified probe on the GlcNAc-neoglycoconjugate SAM at bond rupture forces of 47 ± 15 pN. Additionally, less frequent GlcNAc-specific unbinding events were detected at higher forces (120 ± 20 pN) which are believed to originate from simultaneous detachment of multiple binding sites from the SAM surface. SPR measurements confirmed that WGA has higher affinity toward the immobilized GlcNAc-SAM than toward the soluble free monosaccharide. The binding constants obtained for soluble chitinoligosaccharides suggested up to three subsites within one carbohydrate-binding site of the WGA molecule and also provided further evidence of the multivalent binding character of the WGA dimer.

AB - Carbohydrate–protein interactions govern many crucial life processes involved in cell recognition events, but are often difficult to study because the interactions are weak, and multivalent exposure appears to be crucial for their biological function. We have used self-assembled monolayers (SAMs) of neoglycoconjugates as a model system to probe the specific interactions between the lectin wheat germ agglutinin (WGA) and monosaccharides by surface plasmon resonance (SPR) and atomic force microscopy (AFM) force measurements. SAMs presenting N-acetyl-D-glucosamine (GlcNAc) as a neoglycoconjugate were produced on gold surfaces, where the SAM formation was monitored using a quartz crystal microbalance (QCM) and shown to be a very rapid process. In the AFM force measurements WGA was covalently coupled to flexible polyethylene glycol (PEG) molecules at a probe surface using amine coupling. GlcNAc-specific binding events were detected with a WGA-modified probe on the GlcNAc-neoglycoconjugate SAM at bond rupture forces of 47 ± 15 pN. Additionally, less frequent GlcNAc-specific unbinding events were detected at higher forces (120 ± 20 pN) which are believed to originate from simultaneous detachment of multiple binding sites from the SAM surface. SPR measurements confirmed that WGA has higher affinity toward the immobilized GlcNAc-SAM than toward the soluble free monosaccharide. The binding constants obtained for soluble chitinoligosaccharides suggested up to three subsites within one carbohydrate-binding site of the WGA molecule and also provided further evidence of the multivalent binding character of the WGA dimer.

KW - AFM force spectroscopy

KW - protein-carbohydrate interaction

KW - self-assembled monolayer

KW - surface plasmon resonance

KW - wheat germ agglutinin

U2 - 10.1093/glycob/cwp030

DO - 10.1093/glycob/cwp030

M3 - Article

VL - 19

SP - 633

EP - 643

JO - Glycobiology

JF - Glycobiology

SN - 0959-6658

IS - 6

ER -