Abstract
Proper determination of agonist efficacy is indispensable in the evaluation of agonist selectivity and bias to activation of specific signalling pathways. The operational model (OM) of pharmacological agonism is a useful means for achieving this goal. Allosteric ligands bind to receptors at sites that are distinct from those of endogenous agonists that interact with the orthosteric domain on the receptor. An allosteric modulator and an orthosteric agonist bind simultaneously to the receptor to form a ternary complex, where the allosteric modulator affects the binding affinity and operational efficacy of the agonist. Allosteric modulators are an intensively studied group of receptor ligands because of their selectivity and preservation of physiological space–time pattern of the signals they modulate. We analysed the operational model of allosterically-modulated agonism (OMAM) including modulation by allosteric agonists. Similar to OM, several parameters of OMAM are inter-dependent. We derived equations describing mutual relationships among parameters of the functional response and OMAM. We present a workflow for the robust fitting of OMAM to experimental data using derived equations.
Highlights
Proper determination of agonist efficacy is indispensable in the evaluation of agonist selectivity and bias to activation of specific signalling pathways
Allosteric ligands bind to a site that is distinct from the orthosteric site on a receptor
It has been shown that these three parameters ( EMAX, KA, and τA) are inter-dependent and, we proposed a two-step procedure to overcome this p itfall[15]
Summary
Proper determination of agonist efficacy is a cornerstone in the assessment of possible agonist selectivity and signalling bias. The inherent glitch in OM is that the objective parameters (agonist equilibrium dissociation constant K A, its operational efficacy τA and maximal possible response of the system E MAX) describing it are inter-dependent[15] To circumvent this pitfall, we proposed a two-step procedure of fitting of OM to experimental data. Operational efficacies τA, τB, equilibrium dissociation constants K A and K B, the factor of binding cooperativity α and the maximal response of the system EMAX should be determined before fitting Eq (14) to experimental data (Fig. 10). Eq (14) with K A, KB, τA and τB fixed to values predetermined in functional experiments (Fig. 13) was fitted to the concentration–response curves (global fit) to determine confidence intervals of cooperativity factors α and β. OMAM Eq (5) possesses five and Eq (14) possesses six interdependent parameters making their direct fitting to the experimental data impossible
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