Friday, 29 March 2024

MTA-SZTE Biomimetic Systems Research Group

Principal investigator: Gábor Tóth

Characterization of (model) receptor-ligand and protein-drug molecule interactions by using two-dimensional assays.

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Better understanding of numerous receptor/protein-biomolecule bindings is in the focus of extensive research. Contrary to the (radio)-labelled techniques in recent years a number of “label-free” techniques have been developed to report biomolecular interactions. Surface plasmon resonance (SPR) is label-free technique and capable of measuring real-time quantitative binding affinities and kinetics for proteins interacting with biomolecules using relatively small quantities of materials and has potential to be medium-throughput. The conventional SPR technique requires one binding component to be immobilised on a sensor chip while the other binding component in solution is flowed over the sensor surface; a binding interaction is detected using an optical method that measures small changes in refractive index at the sensor surface. If the small molecules bind to the immobilised proteins, there is an association phase during which binding sites become occupied and the positive slope of SPR curve can be used to measure the rate of association (ka). When steady-state is achieved the RIU (refractive index unit) value corresponds to the changed final critical angle (angle modulated type) or the final wavelength of maximal plasmonic loss (wavelength modulated type). This maximum RIU value relates to the concentrations of immobilised protein and analyte molecules and so can be used to measure the binding affinity (KD=kd/ka). When small molecules are removed from the continuous flow there is a dissociation phase during which binding sites become unoccupied and the (negative slope) of curve can be used to measure the rate of dissociation (kd). Moreover, we can study these biomolecular interactions at different temperatures as well. Using this biosensor assay we can provide not only quantitative and kinetic information we also get thermodynamic state functions of the studied interactions. Our preliminary work was to investigate the interaction between three different glutamate receptor model peptides (GluR1201-230, GluR1231-259 and GluR1270-300) and kynurenic acid (KYNA) by SPR experiments [1-3]. The studied system is specifically relevant in the neuroscience.


References

[1] E. Csapó, Z. Majláth, Á. Juhász, B. Roósz, A. Hetényi, J. Tajti, G.K. Tóth, L. Vécsei, I. Dékány: Coll. Surf. B, 123 (2014) 924-929.

[2] E. Csapó, F. Bogár, Á. Juhász, D. Sebők, J. Szolomájer, Zs. Majláth, G.K. Tóth, L. Vécsei, I. Dékány: Coll. Surf. B, 133 (2015) 66-72.

[3] Á. Juhász, E. Csapó, D. Ungor, G. K. Tóth, L. Vécsei, I. Dékány: J. Phys Chem B, 120 (2016) 7844-7850.