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Supplementary material Dimer-based model for heptaspanning membrane receptors Rafael Franco, Vicent Casadó, Josefa Mallol, Sergi Ferré, Kjell Fuxe, Antonio.

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Presentation on theme: "Supplementary material Dimer-based model for heptaspanning membrane receptors Rafael Franco, Vicent Casadó, Josefa Mallol, Sergi Ferré, Kjell Fuxe, Antonio."— Presentation transcript:

1 Supplementary material Dimer-based model for heptaspanning membrane receptors Rafael Franco, Vicent Casadó, Josefa Mallol, Sergi Ferré, Kjell Fuxe, Antonio Cortés, Francisco Ciruela, Carmen Lluis and Enric I. Canela Corresponding author: Canela, E.I. (ecanela@ub.edu). Department Bioquímica i Biologia Molecular, Universitat de Barcelona, A. Diagonal, 645. 08028 Barcelona, Spain

2 A+A+(RR)A+A(RR) Scheme of the model Inactive Active Vacant Occupied Constitutive A(RR)A A+A+(RR)*A+A(RR)*A(RR)*A L   L  L KK KK   K   K DIMER OPERATING UNIT

3 Constants List of the equilibrium constants for the two-state dimer model ParameterDescription Definition a KEquilibrium association constant of A to RR LEquilibrium receptor isomerization constant  Intrinsic efficacy of A in relation to the binding to unoccupied receptors  Intrinsic efficacy of A in relation to the binding to single-occupied receptors  Binding cooperativity between first and second A molecule: ratio of affinity of A for A(RR) and (RR) [µK being the equilibrium association constant of A to A(RR)] a In this symmetric dimer model [A(RR)] refers to the concentration of dimer with A bound, irrespective of whether A is bound to one site or the other.

4 Ligand binding I. Functions The saturation function is a 2:2 function The ligand-binding function

5 Ligand binding II. Cooperativity analysis Saturation function Reference saturation function K is the average association constant It corresponds to a theoretical non- cooperative binding of A to a dimer

6 Ligand binding III. Cooperativity analysis If Positive cooperativity Negative cooperativity Non-cooperativity. Occurs when If

7 .  p 2 gives an idea of the affinity  positive cooperativity occurs when p 1 < 2·[A] 50  negative cooperativity occurs when p 1 > 2·[A] 50  p 1 = 2·[A] 50 gives non-cooperativity where being Ligand binding IV. Fitting data to the model Rearranging and defining p 1 and p 2

8 Signalling analysis. Functional activity (at zero time) Constitutive activity The function Functional activity (at zero time) at ligand-saturating concentration is proportional to (  – 1)

9 Previously described models

10 del Castillo and Katz model A+RARAR* VacantOccupied ActiveInactive KK LL del Castillo J. and Katz B. (1957) A comparison of acetylcholine and stable depolarizing agents. Proc. R. Soc. Lond. B Biol. Sci. 146, 362  368

11 A+G+R AGR A+GR* AGR* Ternary complex model InactiveActive Vacant Occupied Constitutive KK MM MM   K De Lean A. et al. (1980) A ternary complex model explains the agonist-specific binding properties of the adenylate cyclase-coupled  -adrenergic receptor. J. Biol. Chem. 255, 7108  7117

12 A+G+R G+AR A+G+R* G+AR* Extended ternary complex model Inactive Active Vacant Occupied Constitutive A+R*G AR*G K   K  K LL MM   L   M Samama P. et al. (1993) A mutation-induced activated state of the  2 -adrenergic receptor. Extending the ternary complex model. J. Biol. Chem. 268, 4625  4636

13 A+R+GA+R*+G Cubic ternary complex model InactiveActiveVacantOccupiedConstitutive AR+GAR*+G A+RGA+R*G ARGAR*G K KK LL   L MM MM KK MM LL   L   M  K Weiss J.M. et al. (1996) The cubic ternary complex receptor  occupancy model I. Model description. J. Theor. Biol. 178, 151–167 Weiss J.M. et al. (1996) The cubic ternary complex receptor  occupancy model II. Understanding apparent affinity. J. Theor. Biol. 178, 169–182 Weiss J.M. et al. (1996) The cubic ternary complex receptor  occupancy model III. Resurrecting efficacy. J. Theor. Biol. 181, 381–397

14 A+B+R AR+B A+RB ARB Ternary complex model of allosteric modulation InactiveActive Vacant Occupied Constitutive KK JJ   J   K Tuček S. and Proška J. (1995) Allosteric modulation of muscarinic acetylcholine receptors. Trends Pharmacol. Sci. 16, 205–211

15 A+R+BA+R*+B The allosteric two-state model Inactive Active Vacant Occupied Constitutive AR+BAR*+B A+RBA+R*B ARBAR*B K KK LL   L MM MM KK MM LL   L   M  K Hall D.A. (2000) Modeling the functional effects of allosteric modulators at pharmacological receptors: an extension of the two-state model of receptor activation. Mol. Pharmacol. 58, 1412  1423

16 Two independent sites model A+RAR K1K1 A+R*AR* K2K2 Inactive Active Vacant Occupied Constitutive

17 A+R A+R* AR AR* K2K2 K1K1 K4K4 K3K3 The cluster-arranged cooperative model Franco R. et al. (1996) The cluster-arranged cooperative model: a model that accounts for the kinetics of binding to A 1 adenosine receptors. Biochemistry 35, 3007  3015 Y is the cooperativity factor


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