Structures involved in maintaining the inactive state

One of the truly exciting questions within the GPCR field is what structural changes a receptor undergoes during the transduction of extracellular signals. The classical 'ternary complex model' postulates that receptor activation leads to the agonist-promoted formation of an active 'ternary' complex of agonist, receptor, and G protein (de Lean etal. 1980). This model had to be extended in order to account for the fact that many receptors can activate G proteins in the absence of agonist (Lefkowitz et al. 1993). Based on these seminal observations, receptors are assumed to exist in an equilibrium between the inactive state R and the active state R* (see Chapters 3 and 8). In vitro mutagenesis studies with several GPCRs provided compelling evidence for the existence of intramolecular constraining determinants which stabilize the inactive receptor conformation and are described in detail within Chapter 3. Alteration of such intramolecular contact sites can lead to constitutive receptor activation.

Most of the mutations found to be responsible for constitutive receptor activity are located in the C-terminal portion of the i3 loop and within different TMDs. Interhelical salt bridges, as specific structural determinants stabilizing the inactive state, have been identified in rhodopsin (Robinson et al. 1992) and the a1B adrenergic receptor (Porter et al. 1996). In the LHR, a tightly packed hydrophobic cluster and a specific H-bonding network formed between the cytoplasmic portions of TMD5 and TMD6 and the central regions of TMD6 and TMD7 is thought to maintain the inactive receptor conformation. Mutagenesis data also suggest that LHR activation is associated with the disruption of key interhelical side-chain interactions (Lin et al. 1997). The identification of such specific contact sites provides valuable information about the orientation of the different helices relative towards each other. Recent data with constitutive active LHR mutants implicate that in addition to interhelical interactions the inactive conformation of GPCRs is also stabilized by specific intrahelical structures (Schulz etal. 2000a; see Chapter 3).

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