System dependent and independent measurements

Quantification of ligand activity at GPCRs is unique in that GPCRs are societal proteins residing in synoptic systems. Therefore, the activity of ligands on GPCRs can vary with respect to the type of system in which the receptor resides. This immediately differentiates the types of measurements that can be made on GPCRs, namely system dependent and system independent ligand activity. The aim of quantitation of ligand-GPCR activity is to obtain system independent measurements since these can be used to predict effects in other systems, and eventually, in therapeutic settings.

Extended ternary complex model

Extended ternary complex model

(b) Cubic ternary complex model

(b) Cubic ternary complex model

8yaK

[Ri]

Concentration of receptor in the inactive state (i.e. this species does not activate G-proteins)

[Ra]

Concentration of receptor in the active state (this species activates G-proteins)

[G]

Concentration of G-protein in the system

Ka

Equilibrium association constant for agonist and receptor

Kg

Equilibrium association constant for receptor and G-protein

L

Allosteric constant denoting the ratio of receptor in the active versus inactive state (L = [Ra] / [Ri])

Y

Factor defining the differential affinity of the receptor for G-proteins when the receptor is ligand bound. In the CTC model y defines effect of ligand on G-protein binding to inactive state receptor

a

Factor defining the differential affinity of the ligand for active versus inactive state Also, the effect of ligand binding on receptor activation

ß

Factor defining the differential affinity of the receptor for G-protein when the receptor is in the active state

5

Factor defining the synergy produced by simultaneous ligand binding on the interaction of the G-protein with the receptor

Fig. 8.1 Two widely applied models of GPCR systems; the extended ternary complex model (ETC model; Samama et al. 1993) and the cubic ternary complex model (CTC model, Wiess et al. 1996a ,b,c).

There are two generally used models, based upon linkage theory (Wyman 1965, 1967; Weber 1972,1975; De Lean etal. 1980) for GPCR behaviour. These are the extended ternary complex (ETC) model (Samama et al. 1993) and the cubic ternary complex (CTC) model (Weiss et al. 1996a,b,c); these are shown schematically in Fig. 8.1. The latter is thermody-namically more complete but also more cumbersome version ofthe former. There is evidence to show that some receptor types require the more thermodynamic completeness and are thus more appropriately modelled with the CTC model (Kenakin et al. 2000). Expressions using both models will be given in this chapter. Figure 8.1 indicates the meaning of the various terms. This chapter will discuss methods attempting to measure system independent estimates of affinity (Ka) and efficacy (a, y, 8).

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