REAL-TIME COMPETITIVE KINETIC-ANALYSIS OF INTERACTIONS BETWEEN LOW-MOLECULAR-WEIGHT LIGANDS IN SOLUTION AND SURFACE-IMMOBILIZED RECEPTORS

With surface plasmon resonance detection it is possible to measure the binding kinetics between a macromolecule in solution and a receptor i mmobilized on a sensor surface. The detector response is proportional to the mass of the analyte that binds to the surface, and therefore, a direct observation of a low-molecular-weight (lmw) analyte (<5000 Da) interacting with its immobilized binding partner is normally not poss ible. I describe here a competitive approach in which a lmw analyte an d a high-molecular-weight analyte react at the same time with the immo bilized receptor. Using this approach it is possible to extend kinetic analysis to lmw analyte-receptor interactions. A qualitative analysis allows rapid affinity ranking of different lmw analytes interacting w ith the same receptor, and a quantitative analysis of binding data all ows the calculation of rate constants for the lmw analyte-receptor int eraction. The competitive kinetics approach may therefore be used as a n alternative to other affinity techniques for the characterization of lmw ligands, for identification of inhibitors, and for drug screening . (C) 1994 Academic Press, Inc.