G-protein binding

G-protein-binding domain on the third intracellular loop... 

All rhodopsin-like G-protein-coupled receptors have a conserved arginine residue at the intracellular end of TM3 and this residue is thought to be crucial for G-protein activation. The third intracellular loop determines the class of G-protein activated by the receptor with the second intracellular loop and C-terminus also influencing G-protein binding in some cases. Four classes of G-protein are known ... 

Using chimaeric receptors it has been shown that swapping the third intracellular loop between receptors also swaps their G-protein selectivity. The G-protein-binding... 

Damaj BB, McColl SR, Neote K, et al. Identification of G-protein binding sites of the human interleukin-8 receptors by functional mapping of the intracellular loops. FASEB J 1996 10(12) 1426-1434. 

As described in more detail below, agonist binding will lead to signaling as well as phosphorylation of Ser and Thr residues, especially, but also, in selected cases, Tyr residues located in intracellular loop-3 and in the C-terminal extension. This post-translational modification alters the affinity of the receptor for various intracellular proteins, including arrestin, which sterically prevents further G-protein binding and functions as an adaptor protein. Also, interaction with other types of scaffolding proteins such as PSD-95-like proteins, is influenced by the phosphorylation state of the receptor. 

The most common second messenger activated by protein/peptide hormones and catecholamines is cyclic adenosine monophosphate (cAMP). The pathway by which cAMP is formed and alters cellular function is illustrated in Figure 10.1. The process begins when the hormone binds to its receptor. These receptors are quite large and span the plasma membrane. On the cytoplasmic surface of the membrane, the receptor is associated with a G protein that serves as the transducer molecule. In other words, the G protein acts as an intermediary between the receptor and the second messengers that will alter cellular activity. These proteins are referred to as G proteins because they bind with guanosine nucleotides. In an unstimulated cell, the inactive G protein binds guanosine diphosphate (GDP). When the hormone... 

This transmembrane signaling system involves a complex consisting of several functional proteins (Figure 7) stimulatory (e.g. P-adrenergic, dopamine Dp serotonin, vasopressin) [124] and inhibitory (e.g. a2-adrenergic, dopamine D2, opiod, and muscarinic) [125] receptors, stimulatory (Gs) and inhibitory (G ) G-proteins, and the catalytic protein, adenylate cyclase. On stimulation of a receptor, an associated G-protein binds GTP and the resulting receptor/G-protein/GTP complex then activates, or inhibits, adenylate cyclase in the catalysis of the synthesis... 

Alves, I. D. Salamon, Z. Varga, E. Yamamura, H. I. Tollin, G. Hruby, V. J., Direct observation of G protein binding to the human 8 opioid receptor using plasmon waveguide resonance spectroscopy, J. Biol. Chem. 2003, 278,48890 48897... 

A key feature of both vertebrate and invertebrate opsin molecules is their ability to interact with a G protein, typically transducin, to initiate phototransduction (Ebrey Koutalos 2001). The third cytoplasmic loop that connects a-hehces V and VI contributes to the G protein binding and activation and is highly conserved amongst the rod and cone opsins (Fig. 1, Table 2). This conservation extends to the P opsin family. However, the third cytoplasmic loop of the VA... 

Fig. 3. WEB representation ot GRiFFiN, which aiiows the user to predict G-protein binding seiectivity by entering GPCR sequence and iigand moiecuiar weight.

Hamm, H. E., Deretic, D., Arendt, A., Hargrave, P. A., Koenig, B., and Hofmann, K. P. (1988). Site of G protein binding to rhodopsin mapped with synthetic peptides from the a subunit. Science 241, 832-835. 

A measurement system that is able to quantitatively determine the interactions of receptor and G protein has the potential for more detailed testing of ternary complex models. The soluble receptor systems, ([l AR and FPR) described in Section II, allow for the direct and quantitative evaluation of receptor and G protein interactions (Simons et al, 2003, 2004). Soluble receptors allow access to both the extracellular ligandbinding site and the intracellular G protein-binding site of the receptor. As the site densities on the particles are typically lower than those that support rebinding (Goldstein et al, 1989), simple three-dimensional concentrations are appropriate for the components. Thus, by applying molar units for all the reaction components in the definitions listed in Fig. 2A, the units for the equilibrium dissociation constants are molar, not moles per square meter as for membrane-bound receptor interactions. These assemblies are also suitable for kinetic analysis of ternary complex disassembly. 

G-proteins exist either in an active or an inactive state, depending on the guanylate nucleotide that is bound. In the inactive state, G-protein binds to GDP. In the active state, GTP is bound to the G-pro-tein. G-proteins have an intrinsic GTPase activity, which converts bound GTP to GDP. Hydrolysis of GTP by the G-protein converts the G-protein back to an inactive state. Thus the cycle of the G-protein is as follows ... 

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