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Unlike oxygen, which causes graded allosteric changes in the activity of hemoglobin, some other allosteric effectors act as switches, turning the activity of many different proteins on or off by binding to them noncovalently. Two important allosteric switches that we will encounter many times throughout this book, especially in the context of cell signaling pathways (see Chapters 15 and 16), are Ca²⁺ and GTP.

Ca²⁺/Calmodulin-Mediated Switching The concentration of Ca²⁺ that is free in the cytosol (not bound to molecules other than water) is kept very low (~10⁻⁷ M) by specialized membrane transport proteins that continually pump excess Ca²⁺ out of the cytosol (see Chapters 11 and 15). However, as we will learn in Chapters 11 and 15, the cytosolic Ca²⁺ concentration can increase tenfold to a hundredfold when Ca²⁺-permeable channels in the cell-surface membranes open and allow extracellular Ca²⁺ to flow into the cell. This rise in cytosolic Ca²⁺ is sensed by specialized Ca²⁺-binding proteins, which alter cellular behavior by turning the activities of other proteins on or off. The importance of extracellular Ca²⁺ for cell activity was first documented by S. Ringer in 1883, when he discovered that isolated rat hearts suspended in an NaCl solution made with “hard” (Ca²⁺-rich) London tap water contracted beautifully, whereas they beat poorly and stopped quickly if distilled, Ca²⁺-depleted, water was used.

Many Ca²⁺-binding proteins bind Ca²⁺ using the EF hand/helix-loop-helix structural motif discussed earlier (see Figure 3-10b). A well-studied EF hand protein, calmodulin, is found in all eukaryotic cells, where it may exist as an individual monomeric protein or as a subunit of a multimeric protein. This dumbbell-shaped molecule contains four Ca²⁺-binding EF hands with K_ds of about 10⁻⁶ M. The binding of Ca²⁺ to calmodulin causes a conformational change that permits Ca²⁺/calmodulin to bind to conserved sequences in various target proteins (Figure 3-33), thereby switching their activities on or off. Calmodulin and similar EF hand proteins thus function as switch proteins, acting in concert with changes in Ca²⁺ levels to modulate the activity of other proteins.

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Switching Mediated by Guanine Nucleotide–Binding Proteins Another group of intracellular switch proteins constitutes the GTPase superfamily. As the name suggests, these proteins are enzymes—GTPases—that can hydrolyze GTP (guanosine triphosphate) to GDP (guanosine diphosphate). They include the monomeric Ras protein (whose structure is shown in Figure 3-9, with bound GDP shown in blue) and the G_α subunit of the trimeric G proteins, both discussed at length in Chapters 15 and 16. Both Ras and G_α can bind to the plasma membrane, function in cell signaling, and play key roles in cell proliferation and differentiation. Other members of the GTPase superfamily function in protein synthesis, the transport of proteins between the nucleus and the cytoplasm, the formation of coated vesicles and their fusion with target membranes, and rearrangements of the actin cytoskeleton. Some GTPase proteins have a covalently attached lipid chain (see Figure 7-19) that mediates their binding to membranes. We examine the roles of various GTPase switch proteins in regulating intracellular signaling and other processes in several later chapters.

All the GTPase switch proteins exist in two forms, or conformations (Figure 3-34): (1) an active (“on”) form with bound GTP, which can influence the activity of specific target proteins to which they bind, and (2) an inactive (“off”) form with bound GDP. The switch is turned on—that is, the conformation of the protein changes from inactive to active—when a GTP molecule replaces a bound GDP in the inactive conformation. The switch is turned off when the relatively slow GTPase activity of the protein hydrolyzes bound GTP, converting it to GDP and leading the conformation to change to the inactive form. The amount of time any given GTPase switch remains in the active, GTP-bound form depends on the rate of its GTPase activity. Thus the GTPase activity acts as a timer to control this switch. Cells contain a variety of proteins that can modulate the baseline (or intrinsic) rate of GTPase activity for any given GTPase switch and so can control how long the switch remains on. For example, GTPase-activating proteins, or GAPs, increase the rate of GTPase activity, thus reducing the time the GTPase is in the active form. Cells also have specific proteins whose function is to regulate the conversion of inactive GTPases to active ones—that is, to turn the switch on—by mediating the replacement of bound GDP with GTP (GDP/GTP exchange). These proteins are called guanine nucleotide exchange factors, or GEFs. GTPases with lipid anchors are also regulated by proteins called guanine nucleotide dissociation inhibitors (GDIs) that bind to the lipid chain and thus influence interactions with cellular membranes.