Kinetic Binding Analysis of Biomolecular Interactions Using Surface Plasmon Resonance System

Abstract

Surface plasmon resonance (SPR) technology is a versatile and sensitive tool widely used for kinetic studies of interactions between unlabelled biomolecules in real time. The interactions of 4-carboxy benzene sulfonamide (4-CBS) inhibitor, with the enzyme carbonic anhydrase II (CA II) and Interleukin 2 with its antibody (IL 2 antibody) were determined at different concentrations by injecting at a flow rate of 100 μl.min-1 for 60s (100 μl) for a 1 min association phase, which was followed by a 2 min dissociation phase. Langmuir model yielded the rate constants ka and kd , which were of the order of 1.58E+3 M-1. s-1 and 0.04 s-1, respectively, and equilibrium dissociation constant was 2.45E-7 M. Automatic artifact removal, using software options and double referencing (by injection of running buffer to nullify non-specific interactions), further improved the quality of response. Langmuir model yielded the rate constants ka and kd , which were of the order of 2.79E+5 M-1.s-1 and 1.43E-4 s-1, respectively, and equilibrium dissociation constant was 5.17E-7 M. The results illustrate the potential of such kinetic analysis in disease management, drug discovery and for detection of pathogens, toxins, pesticides and antibiotic residues based on either enzyme inhibition or antigen-antibody binding