Dissociative GTP

Fig. 2. Minimal components of the G protein-coupling cycle in a schematic chemotaxis pathway. (A) Chemotaxis G protein-coupled receptors reside in the plasma membrane associated with specific heterotrimeric G proteins. (B) Upon binding of a chemoattractant such as fMLF, the receptor catalyzes the exchange of GTP for GDP in the a subunit of the G protein, thereby dissociating the GTP-bound a subunit from the (3 subunit complex. Chemotaxis GPCRs typically utilize the a isoform of the a subunit. (C) Subsequently the dissociated GTP-bound a subunit and the j3 subunit complex each dock to effectors elsewhere in the cell. (D) The a subunit possesses intrinsic GTPase activity that hydrolyzes the bound GTP to GDP, thereby regenerating the GDP-bound Oj subunit that reassociates with the / 7 subunit complex and with the receptor. Bordetella pertussis toxin covalently and specifically modifies the isoform of the a subunit and prevents its association with receptor (see text for additional discussion and references).

Fig. 10. The receptor—G-protein sequence. An activated receptor interacts with the trimeric GDP-ligated receptor to cause an interchange of GDP by GTP and dissociation into the activated Ga—GTP (left) and G y (right) subunits. These then interact with a variety of effectors. The purpose of the activated...

FIGURE 15.21 Hormone (H) binding to its receptor (R) creates a hormone receptor complex (H R) that catalyzes GDP-GTP exchange on the o -subunit of the heterotrimer G protein (G ), replacing GDP with GTP. The G -subunit with GTP bound dissociates from the /37-subunits and binds to adenylyl cyclase (AC). AC becomes active upon association with G GTP and catalyzes the formation of cAMP from ATP. With time, the intrinsic GTPase activity of the G -subunit hydrolyzes the bound GTP, forming GDP this leads to dissociation of G GDP from AC, reassociation of G with the /Sy subunits, and cessation of AC activity. AC and the hormone receptor H are integral plasma membrane proteins G and G are membrane-anchored proteins. 

Stimulation and inhibition of the enzyme by the GPCR-G-protein cycle occur by analogous mechanisms. Agonists induce hormone receptors to increase a Ga-GDP-GTP exchange and subsequent Ga 3y dissociation (GDP-a py + GTP GTP-ax + [3y + GDP) (Fig. 4). Consequently, agents that affect either the dissociation of either G or Gs, or the association of their respective as, a , or (3y subunits with adenylyl cyclase could affect rates of cAMP formation in enzyme preparations or in intact cells and tissues. There are several important examples. Gas is stably activated by poorly hydrolyzable analogs of GTP, e.g. GTPyS... 

In general, the receptor-G-proteins complexes exchange bound GDP for GTP. In turn, the two, smaller subunits of the G-protein components of these complexes are released and the receptor protein dissociates. The remaining G-protein GTP complex then complexes with and activates a specific enzyme. It is very significant to note that G-proteins therefore have at least three specific binding sites (a) for nucleotides, (b) for a receptor protein, and (c) an effector protein. 

The binding of an agonist to a receptor promotes the formation of a ternary LRG complex and an exchange of guanosine-5 -triphosphate (GTP) for guanosine-5 -diphosphate (GDP) on the G protein. The ternary complex is thus activated, promoting dissociation of LR from G and G into its subunits G and... 

The physiologic LRG interaction actually involves at least five types of components ligand, receptor, intact G or G protein subunits (a and Py), GTP (which promotes LRG dissociation and activation), and GDP (which promotes LR dissociation from G but not G dissociation into its subunits). Little about the dynamics of these processes is known. The LRG states may be visualized, in shorthand, as a 3-dimensional array (Figure 1). A large number of questions concerning LRG states remain to be addressed ... 

Figure 3. The binding and dissociation of FU EP and receptor on permeabilized neutrophils at 37 C The data are plotted as the specific binding (pmoles FLPEP bound/lO cells) on a log plot versus time. 10 permeabilized cells/mL were exposed to 1 nM FLPEP at time 0. After IS or 120 s, antibody to fluorescein was added to each sample. 60 seconds later, 10- M GTP yS was added. (Reproduced with permission from reference 22. Copyright 1987 Journal of Biological Chemistry.)...

Initiation of protein synthesis requires that an mRNA molecule be selected for translation by a ribosome. Once the mRNA binds to the ribosome, the latter finds the correct reading frame on the mRNA, and translation begins. This process involves tRNA, rRNA, mRNA, and at least ten eukaryotic initiation factors (elFs), some of which have multiple (three to eight) subunits. Also involved are GTP, ATP, and amino acids. Initiation can be divided into four steps (1) dissociation of the ribosome into its 40S and 60S subunits (2) binding of a ternary complex consisting of met-tRNAf GTP, and eIF-2 to the 40S ribosome to form a preinitiation complex (3) binding of mRNA to the 40S preinitiation complex to form a 43S initiation complex and (4) combination of the 43S initiation complex with the 60S ribosomal subunit to form the SOS initiation complex. 

Step 4 Coat disassembly (involving dissociation of ARF and coatomer shell) follows hydrolysis of bound GTP uncoating is necessary for fusion to occur. 

The structure of the G-protein is heterotrimeric. Following activation by GTP binding, the trirner dissociates into its a and [iv subunits, each of which may elicit cell responses. 

Binding of GTP promotes a disordering of the carboxyl- and amino-termini of the G-protein a subunit, with two parallel consequences the GTP-bound a subunit dissociates... 

The terminal (y) phosphate of GTP is hydrolyzed by the GTPase activity of the G-protein a subunit, leaving GDP bound instead. This reverses the conformational change in step 5 and allows the a subunit to dissociate from the effector and reassociate with the py subunit. The reassociation will also reverse Py-effector interaction because Ga-GDP effectively competes with the effector for Py-binding. Though of fairly high affinity (e.g.,... 

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