Journal
JOURNAL OF LEUKOCYTE BIOLOGY
Volume 90, Issue 2, Pages 399-408Publisher
FEDERATION AMER SOC EXP BIOL
DOI: 10.1189/jlb.1010565
Keywords
chemokine; biosensor; glycosaminoglycan; affinity constant; chemokine receptor
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Funding
- Spanish Ministry of Health FISS
- Spanish Ministry of Science and Innovation [SAF 2008-03388, SAF 2008-00908]
- FISS [RD08/0075/0010]
- European Union [LSHB-CT-2005-518167, 223404]
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Use of SPR-based biosensors is an established method for measuring molecular interactions. Their application to the study of GPCRs is nonetheless limited to detergent-solubilized receptors that can then be reconstituted into a lipid environment. Using the chemokine receptor CXCR4 and its specific ligand CXCL12, we outline here a highly reproducible biosensor method based on receptor presentation on the surface of lentiviral particles; the approach is simple and does not require the use of antibodies to achieve correct receptor orientation on the sensorchip surface. We measured the kinetic parameters of CXCR4/CXCL12 binding in a single step and in real time and evaluated the effect of GAG presentation of chemokines on this interaction. The data indicate that at low concentrations, soluble heparin modulates CXCR4/CXCL12 interaction and at high concentrations, abrogates binding. These observations suggest that in addition to their known role in modulating local chemokine availability, GAG affect the receptor/ligand interaction, although their influence on affinity parameters is very limited. The method will also be useful for quantifying these biomarkers in biological fluids and for the development of high-throughput screening for their antagonists. J. Leukoc. Biol. 90: 399-408; 2011.
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