Like the G_α subunits in trimeric G proteins discussed in Chapter 15, monomeric Ras proteins are “switch” proteins that alternate between an active “on” state with a bound GTP and an inactive “off” state with a bound GDP (see Figures 15-4 and 15-5 to review this concept). Unlike trimeric G proteins, Ras is not directly linked to cell-surface receptors. Like that of trimeric G proteins, Ras activity is regulated by several other proteins. Ras activation is accelerated by a guanine nucleotide exchange factor (GEF), which binds to the Ras·GDP complex, causing dissociation of the bound GDP (see Figure 15-4). As with other G proteins, GTP binds spontaneously to “empty” Ras molecules, forming the active Ras·GTP.

Ras (~170 amino acids) is smaller than G_α proteins (~300 amino acids), but the GTP-binding domains of the two protein types have a similar structure (see Figure 15-5 to review the structure of Ras). Hydrolysis of the bound GTP to GDP deactivates Ras. Structural and biochemical studies show that G_α proteins contain a GTPase-activating protein (GAP) domain that increases the intrinsic rate of GTP hydrolysis. Because this domain is not present in Ras, that protein has an intrinsically slower rate of GTP hydrolysis. Thus the average lifetime of a GTP bound to Ras is about 1 minute, which is much longer than the average lifetime of a G_α·GTP complex. Because the intrinsic GTPase activity of Ras·GTP is low compared with that of G_α·GTP, Ras·GTP requires the assistance of another protein, a GTPase-activating protein (GAP), to deactivate it. Binding of a GAP to Ras·GTP accelerates the intrinsic GTPase activity of Ras by more than a hundredfold; the actual hydrolysis of GTP to GDP and P_i is catalyzed by amino acids from both Ras and GAP. In particular, insertion of one of GAP’s arginine side chains into the Ras active site stabilizes an intermediate in the hydrolysis reaction.

The first indication that Ras functions downstream from RTKs in a common signaling pathway came from experiments in which cultured fibroblast cells were induced to proliferate by treatment with a mixture of two protein hormones that activate RTKs: PDGF and EGF. Microinjection of anti-Ras antibodies into these cells blocked cell proliferation. Conversely, injection of Ras^D, a constitutively active mutant Ras protein that hydrolyzes GTP very inefficiently and thus persists in the active state, caused the cells to proliferate in the absence of the growth factors. These findings are consistent with studies, using the pull-down assay method detailed in Figure 15-11, in which addition of FGF to fibroblasts led to a rapid increase in the proportion of Ras present in the GTP-bound active form. However, as we will see, an activated RTK (or cytokine receptor) cannot directly activate Ras. Instead, other proteins must first be recruited to the activated receptor, where they serve as adapters.