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Supplementary material Dimer-based model for heptaspanning membrane receptors

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

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Supplementary material Dimer-based model for heptaspanning membrane receptors

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  1. Supplementary materialDimer-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. Scheme of the model DIMER OPERATING UNIT Inactive Vacant K mK A+A+(RR) A+A(RR) A(RR)A a L L qa L a K mq K A+A+(RR)* A+A(RR)* A(RR)*A Constitutive Occupied Active

  3. Constants List of the equilibrium constants for the two-state dimer model aIn 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 ligand-binding function The saturation function is a 2:2 function

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

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

  7. Ligand binding IV. Fitting data to the model Rearranging and defining p1 and p2 where being . • p2 gives an idea of the affinity • positive cooperativity occurs when p1 < 2·[A]50 • negative cooperativity occurs when p1 > 2·[A]50 • p1 = 2·[A]50 gives non-cooperativity

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

  9. Previously described models

  10. del Castillo and Katz model Inactive Active K L A+R AR AR* Vacant Occupied 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. Ternary complex model Constitutive Vacant M A+G+R A+GR* Inactive Active K a K aM AGR AGR* Occupied De Lean A. et al. (1980) A ternary complex model explains the agonist-specific binding properties of the adenylate cyclase-coupled b-adrenergic receptor. J. Biol. Chem. 255, 71087117

  12. Extended ternary complex model Vacant Constitutive L mM A+G+R A+G+R* A+R*G a K K qa K a L mq M G+AR G+AR* AR*G Occupied Active Inactive Samama P. et al. (1993) A mutation-induced activated state of the b2-adrenergic receptor. Extending the ternary complex model. J. Biol. Chem. 268, 46254636

  13. Cubic ternary complex model abd L ARG AR*G 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 gM bgd M gK agdK a L AR+G AR*+G bL A+RG A+R*G aK K M bM L A+R+G A+R*+G Vacant Occupied Active Inactive Constitutive

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

  15. The allosteric two-state model abd L ARB AR*B gM Vacant bgd M gK agdK Occupied a L AR+B AR*+B Inactive bL A+RB A+R*B Active aK K M bM Constitutive L A+R+B A+R*+B 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 Vacant K1 A+R AR Occupied Inactive K2 A+R* AR* Active Constitutive

  17. The cluster-arranged cooperative model K1 A+R AR K2 K4 K3 A+R* AR* Y is the cooperativity factor Franco R. et al. (1996) The cluster-arranged cooperative model: a model that accounts for the kinetics of binding to A1 adenosine receptors. Biochemistry 35, 30073015

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