Kinetic measurement of the interaction between an oligosaccharide and lectins by a biosensor based on surface plasmon resonance

Abstract

Kinetic measurements of the interaction between an oligosaccharide and various lectins were performed using a biosensor based on surface plasmon resonance (SPR). A glycopeptide, prepared from asialofetuin and having a nearly homogeneous N-linked sugar chain, was immobilized on the surface of a sensor chip via the amino groups of its peptide moiety. The interactions of this bound glycopeptide with six lectins [Sambucus sieboldiana lectin, Maackia amurensis lectin, Aleuria uuruntia lectin, Ricinus communis agglutinin-1 20 (RCA,,,), Datura stramonium lectin (DSA) and Phaseolus vulgaris leukoagglutinating lectin] were monitored in real-time with the change in the SPR response. Of these lectins, only RCA,,, and DSA showed an increase in the SPR response, indicating that these two lectins bound specifically to the immobilized glycopeptide. The other lectins did not show any significant changes in the SPR response. These results are in good agreement with the binding specificity previously demonstrated with affinity chromatography. The association-rate constant (k,,,) and the dissociation-rate constant (k,,,,) for the glycopeptide-RCA,,, interaction were 3.4X lo5 M-' s-' and 2.1 X lop3 s-', respectively. The k,,, and kdlSs determined for DSA were S.7X105 M-' s-' and 1.3X 10-3 sp', respectively. Furthermore, the relative binding molar ratio to the glycopeptide was three times higher for RCA,,, than for DSA, suggesting that this sugar chain possesses three binding sites for RCA,,, and one for DSA. These parameters are expected to provide useful information for defining the interaction between oligosaccharides and lectins.Many aspects of the interactions between lectins and glycoproteins have been studied, especially regarding the recognition of molecules. These interactions have been used for the detection of the oligosaccharides of glycoproteins. In conventional methods, the recognition and adsorption properties of lectins with the specific sugar residues of glycoproteins are used for chromatographic purification and electrophoretic analytical techniques [l-31. However, it is not easy to elucidate the reaction mechanisms. Furthermore, for kinetic studies of the interactions between lectins and glycoproteins, it is usually necessary to label the molecules, either lectins or carbohydrates [4, 51. This has been a time-consuming step in these studies. The method introduced here requires no labeling, and the interaction phenomena of biological molecules can be monitored directly by the surface plasmon resonance (SPR).The phenomena of SPR was studied by Otto [6] and Kretschman and Raether [7], and it was used as a chemical Correspondence to Y. Shinohara,

Kinetics and the Mechanism of Interaction of the Endoplasmic Reticulum Chaperone, Calreticulin, with Monoglucosylated (Glc1Man9GlcNAc2) Substrate

“…5a); thus, the nature of the reaction is unaffected by immobilization of either of the reactants. Studies on interaction between oligosaccharide and plant lectins and C-type animal lectins by SPR showed that the binding affinity values were in good agreement with those previously determined by affinity chromatography and Scatchard analysis (41,42). Moreover, the rate constants for the association of Ricinus communis agglutinin as determined by temperature jump spectroscopy and the SPR method are also found in perfect agreement (42,43).…”

SPR Studies of Carbohydrate–Protein Interactions: Signal Enhancement of Low‐Molecular‐Mass Analytes by Organoplatinum(II)‐Labeling

“…Nevertheless, since the SPR response is proportional to the accumulation of mass on the sensor surface, a serious constraint imposed by this technique concerns the dimensions of the molecules to be employed as analytes. [7] In recent years, several groups have focused on SPR as an emerging technique to detect protein-carbohydrate interactions, [8][9][10][11][12][13][14][15][16][17][18][19][20] key steps in many biological events. [21,22] SPR studies of these biological events are hampered by the low availability of high-molecular-mass oligosaccharides and by the weakness of protein-carbohydrate interactions.…”

Fluorescence Labeling of Oligosaccharides Useful in the Determination of Molecular Interactions

“…6 Although this approach using ABEE was also useful in structural analyses on HPLC and in measurements of lectin-carbohydrate interaction analysis in ELISA, pyrene label surpassed ABEE by about ten times in sensitivity. The immobilization methods used by other groups for the oligosaccharides include covalent coupling to modified surface of plates, 6,8 the binding of biotinylated oligosaccharides to avidin-coated plates, 9 and the production of neoglycolipids. 10 In these methods, additional or entirely different procedures were necessary to modify the oligosaccharides from those used for other detections.…”