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Related Experiment Videos

Analysis of cell-adhesion molecule interactions using surface plasmon resonance

P A van der Merwe1, A N Barclay

  • 1MRC Cellular Immunology Unit, Sir William Dunn School of Pathology, University of Oxford, UK. vdmerwe@molbiol.ox.ac.uk

Current Opinion in Immunology
|April 1, 1996
PubMed
Summary

Surface plasmon resonance (SPR) biosensors enable the study of weak cell adhesion molecules, like CD2 and CD48. Researchers used SPR to analyze molecular interactions, binding affinities, and potential measurement pitfalls.

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Immunology

Background:

  • Cell adhesion is mediated by weak molecular interactions, posing challenges for biochemical analysis.
  • Real-time optical biosensors, particularly surface plasmon resonance (SPR), are crucial for studying these interactions.

Purpose of the Study:

  • To demonstrate the utility of SPR in analyzing cell adhesion molecule interactions.
  • To investigate the binding characteristics of T cell surface molecule CD2 with its ligand CD48.
  • To explore the application of SPR in analyzing lectin-carbohydrate interactions, such as with CD22.

Main Methods:

  • Utilizing surface plasmon resonance (SPR) biosensors for real-time kinetic and affinity measurements.
  • Employing site-directed mutagenesis to delineate interacting surfaces and quantify residue contributions to binding energy.

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  • Combining SPR with in situ modification of carbohydrates on glycoproteins to assess lectin binding specificity.

    • Main Results:

    • SPR revealed that adhesion molecules like CD2 can exhibit very low affinities (Kd ~100 microM) and rapid dissociation rates (half-lives ≤ 0.2s).
    • SPR combined with mutagenesis elucidated the interacting surfaces of CD2 and CD48, their binding orientation, and the energetic contribution of individual residues.
    • SPR facilitated the analysis of CD22 lectin binding specificity through carbohydrate modification.

    Conclusions:

    • SPR is a powerful technique for characterizing weak molecular interactions relevant to cell adhesion.
    • SPR, coupled with mutagenesis, provides detailed insights into the molecular basis of protein-protein interactions.
    • SPR analysis requires careful consideration of potential pitfalls that can affect measurement accuracy.