Regulatory GTPases

The fimction of eIF-2 is illustrated schematically in Fig. 1.55. eIF-2 belongs to the superfamily of regulatory GTPases (see ch. 5). elF-2 fulfills the task of bringing the methionyl-initiator-tRNA to the 40S subimit of the ribosome. The active eIF-2 GTP form binds the methionyl-initiator-tRNA, associates with the cap structure of the mRNA, then commences to scan along the mRNA. Once an AUG codon is encoimte-red, the boimd GTP is hydrolyzed to GDP, resulting in the dissociation of the... 

Fig. 1. 55. The function of eIF-2 in eucaryotic translation. eIF-2, the initiator protein for the translation is a regulatory GTPase that occurs in an active GTP-form and in an inactive GDP form (see ch. 5). The active eIF-2 GTP forms a complex with the initiator-tRNA, fMet-tRNA "" and the 40S subunit of the ribosome. This complex binds to the cap structure of mRNA to initiate the scanning process. eIF-2 undergoes an activation cycle typical for regulatory GTPases the inactive eIF-2 GDP fom is activated with the assistance of the eIF-2B protein into the active elF-2 GTP form. eIF-2B acts as a G-nucleotide exchange factor in the cycle (see ch. 5).

The most important components of intracellular signal transduction are the protein kinases, protein phosphatases, regulatory GTPases and adapter proteins ... 

The regulatory GTPases function as switches that can exist in an active or inactive form. In the active form the GTPases can transmit signals to downstream components in the signaling chain. In the inactive form signal transmission in repressed. 

Proteins of the GTPase superfamily are found in all plant, bacterial and animal systems. The following examples illustrate the central functions of the regulatory GTPases in the cell. 

This relationship is valid if it can be assumed that the GTP concentration is not limited and that GTP binds very rapidly to the empty form of the GTPase. A special characteristic of the regulatory GTPases is that both rate constants may be regulated by specific proteins. The proportion of GTPase that exists in the active form can be altered by at least three processes ... 

Like all regulatory GTPases, the heterotrimeric G-proteins nm through a cyclical transition between an inactive, GDP-boimd form and an active, GTP-boimd form. Fig. 5.16 sketches the different functional states and the role of the individual subunits. 

Gt,a is made up of two domains, a GTPase domain and a helical domain. The GTPase or G-domain indicates that Gt, is a member of the superfamily of regulatory GTPases. In addition, G, possesses a helical domain, which represents a characteristic feature of the heterotrimeric G-proteins. The nucleotide binding site is in a cleft between the two domains. It is assumed that the presence of the helical domain is the reason that bound nucleotide dissociates only very slowly from transducin and that the activated receptor is therefore necessary to initiate the GDP/GTP exchange. 

Fig. 5.19. GTP and GDP structures of transducin. The Ga,t subunit of transducin possesses—in contrast to Ras protein and to other small regulatory GTPases —an a-hehcal domain that hides and closes the G-nucleotide binding pocket. The conformational changes that accompany the transition from the inactive G t GDP form (a) into the active G t GTP form (b), are restricted to three structural sections that are known as switches I, II and III. Switch I includes the link of the a-helical domain with P2, switch II affects in particular hehx a2, and switch III, the pS—a3 loop. Switch III includes a sequence that is characteristic for the a-subunits of the heterotrimeric G-proteins. The conformational changes of switches II and III affect structural sections that are assumed to be binding sites for the effector molecule adenylyl cyclase (AC) and the y-subunit of cGMP-dependent phosphodiesterase (PDEy), based on mutation experiments and biochemical investigations. MOLSKRIPT representation according to Krauhs, (1991).

Following the discovery of the Ras protein, it was quickly estabhshed that Ras proteins are a family within a large superfamily, known today as the Ras superfamily of monomeric GTPases. Hie members of the superfamUy of Ras proteins are regulatory GTPases of 16-25 kDa, which are active as monomers. 

The lifetime of the active GTP-bound state may be reduced by regulatory GTPase activating proteins. The primary fimction of the GTPase activating proteins (GAP) is to negatively regulate the Ras proteins and Ras-related proteins. 

Table 9.1. Regulatory GTPases and effector proteins of the Ras superfamily of mammals. ...

The Ras protein, as a regulatory GTPase, shows the G domain typical for the superfamily of regulatory GTPases (see Fig. 5.12). The sequence motives characteristic for regulatory GTPases (c 5.3.3) are involved in binding the nucleotide and Mg. Three... 

Oncogenic activation of small regulatory GTPases has been documented many times for the example of the Ras proteins (see 9.2.3). 

Sprinzl, M. (1994) Elongation factor Tu a regulatory GTPase with an integrated effector. Trends Biochem. Sci. 19, 245-250. 

Cassel, D., Levkovitz, H., and Selinger, Z. (1977). The regulatory GTPase cycle of turkey erythrocyte adenylate cyclase./ Cyclic Nucleotide Res. 3, 393-406. 

Goody, R. S. (2003). The significance of the free energy of hydrolysis of GTP for signal-transducing and regulatory GTPases. Biophys. Chem. 100, 535-544. 

Fig. 2. Summary of regulatory GTPase cycle in photoactivation of cGMP-specific phosphodiesterase (PDE) in retinal rod cells. T, transducin (Gt) Rho, rhodopsin Rho, photoactivated Rho. PDE is represented as a heterotrimeric peripheral membrane protein, as is T. This regulatory cycle differs from that in Fig. 1 mainly in that the activation of PDE entails the dissociation of an inhibitory y subunit (PDEy) under the influence of activated Ta-GTP complex leading to formation of intermediary soluble Ta-GTP/PDEy complex. This complex persists until GTP is hydrolyzed to GDP, at which moment the inhibited PDEa/3y heterotrimer reforms. Dark adapted - non-activated - Rho is then required for reassociation of Ta-GDP to T/3y and release of GDP.

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