Principles of cell signaling - UT Southwestern

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39057_ch14_cellbio.qxd 8/28/06 5:11 PM Page 624 ERK2 inactive and active conformations INACTIVE (ERK2) N terminal domain Tyr185 Thr183 C terminal domain ACTIVE (ERK2-P2) Tyr185 Thr183 FIGURE 14.31 The structures of unphosphorylated, inactive MAPK ERK2 and phosphorylated, active ERK2 are compared. ERK2 has a typical protein kinase structure. The smaller N-terminal domain is composed primarily of strands and the larger C-terminal domain is primarily -helical. The active site is formed at the interface of the two domains. The activation loop emerges from the active site and is refolded following phosphorylation of the Tyr and Thr residues, inducing the repositioning of active site residues. ATP (not shown) binds in the interior of the active site; productive binding of protein substrates to the surface of the C-terminal domain is also facilitated by the reorganization of the activation loop. Structures generated from Protein Data Bank files 1ERK and 2ERK. Sensor/ Histidine kinase Response regulator His Asp Ligand P ATP His phosphorylation Two-component signaling systems ADP His Asp P P Transfer of phosphate to Asp: response regulator active His Asp P P H 2 O Response regulator deactivated FIGURE 14.32 The basic two-component system is composed of a signal-activated histidine kinase, referred to as a sensor, and an effector protein, the response regulator, that is activated when it is phosphorylated on an aspartate residue by the sensor. The activity of the response regulator is terminated when the aspartyl-phosphate is hydrolyzed. the protein kinase family. There are unique inserts on the surface of protein kinases that generate specificity in localization, interaction with other regulatory molecules, and recognition of substrates. These landmarks allow both classification and genetic manipulation of protein kinases. Protein kinases have evolved numerous and diverse regulatory mechanisms to complement their number and multiple functions. These mechanisms include allosteric activation and inhibition by lipids, soluble small molecules and other proteins; activating and inhibitory phosphorylation and other covalent modifications, including proteolysis; and binding to scaffolds and adaptors to enhance activity or limit nonspecific activities. Many such inputs may regulate a single protein kinase in a complex combinatoric code. Further, multiple protein kinases that act sequentially, such as in a protein kinase cascade (see Figure 14.38), can create uniquely complex signaling patterns. 14.25 Two-component protein phosphorylation systems are signaling relays Key concepts • Two-component signaling systems are composed of sensor and response regulator components. • Upon receiving a stimulus, sensor components undergo autophosphorylation on a histidine (His) residue. • Transfer of the phosphate to an aspartyl residue on the response regulator serves to activate the regulator. Prokaryotes, plants, and fungi share an alternative mechanism for regulatory phosphorylation and dephosphorylation known as two-component signaling. FIGURE 14.32 shows a typical twocomponent system. In this system, the receptor, referred to as a sensor, responds to a stimulus by catalyzing its own phosphorylation on a His residue. Sensors include chemoattractant receptors in bacteria, a regulator of osmolarity in fungi, light-sensitive proteins, the receptor for the plantripening hormone ethylene, and other receptors for diverse environmental, hormonal, and metabolic signals. The mammalian mitochondrial dehydrogenase kinases are related in sequence to the bacterial histidine kinases, although the mammalian enzymes phosphorylate serine or threonine residues, not histidine. The phosphorylated sensor next transfers its covalently bound phos- 624 CHAPTER 14 Principles of cell signaling
39057_ch14_cellbio.qxd 8/28/06 5:11 PM Page 625 phate to an aspartyl residue on a second protein known as a response regulator. Response regulators initiate cellular responses, usually by binding to other cytoplasmic proteins and allosterically regulating their activities. Although all two-component systems follow this same general pattern, their structures and precise reaction pathways vary enormously. Some two-component systems are composed of only one protein (sensor and response regulator in a single polypeptide chain). Others are composed of a sensor protein and two aspartylphosphorylated proteins, in which the first or the second may display response regulatory activity. Finally, two-component systems usually lack conventional protein phosphatases. Hydrolysis of the aspartyl-phosphate bond may be spontaneous or regulated by the response regulator itself. 14.26 Pharmacological inhibitors of protein kinases may be used to understand and treat disease Key concepts • Protein kinase inhibitors are useful both for signaling research and as drugs. • Protein kinase inhibitors usually bind in the ATP binding site. Many inhibitors have been developed for basic research purposes to explore the functions of protein kinases. The importance of these enzymes in disease processes has also made them targets of drug screening projects yielding inhibitors for many protein kinases. The majority of pharmacological inhibitors of protein kinases compete with ATP binding. Because of the huge number of ATP-binding proteins in a cell, there are inevitable concerns about inhibitor specificity not only with respect to the other protein kinases but also to the other proteins that bind nucleotides. This problem has been mitigated with variable success through chemical library screening, structure-based modification of lead compounds, and inhibitor testing against panels of protein kinases. Many inhibitors with actions on PKA or PKCs, for example, have effects on several other members of the AGC family. Although pharmacological inhibitors with effects on PKA abound, the most selective are derived from the naturally occurring small inhibitory protein known as PKI or the Walsh inhibitor. In vitro and cell-based screens have identified much more selective inhibitors for MAP2Ks in the ERK1/2 pathway. These inhibitors have fewer known protein kinase cross reactivities, probably due to the fact that they do not bind in the ATP site. Among inhibitors that have progressed in the clinic, compounds developed against the EGF receptor and certain other protein tyrosine kinases have had considerable success. 14.27 Phosphoprotein phosphatases reverse the actions of kinases and are independently regulated Key concepts • Phosphoprotein phosphatases reverse the actions of protein kinases. • Phosphoprotein phosphatases may dephosphorylate phosphoserine/threonine, phosphotyrosine, or all three. • Phosphoprotein phosphatase specificity is often achieved through the formation of specific protein complexes. Protein phosphorylation is reversed by phosphoprotein phosphatases. These enzymes display distinct specificities and modes of regulation. Phosphoprotein phosphatases can be considered in two broad groups based on their specificity and sequence relationships: protein-serine/threonine phosphatases and protein-tyrosine phosphatases. Most protein-serine/threonine phosphatases are regulated by association with other proteins. Targeted localization is the major determinant of substrate specificity. Phosphoprotein phosphatase 1 (PP1) associates with a variety of regulatory subunits that specifically direct it to relevant organelles. One subunit (known as the G subunit), for example, specifies association with glycogen particles. The interaction with this subunit is itself regulated by phosphorylation. Small protein inhibitors can suppress PP1 activity. Phosphoprotein phosphatase 2A (PP2A) is composed of a catalytic subunit, a scaffolding subunit, and one of a large number of regulatory subunits. The regulatory subunit modulates activity and localization of the phosphatase. Some viruses alter the behavior of the cells they infect by interfering with phosphatase activity. For example, cells transformed with the SV40 virus express a viral protein known as small t anti- 14.27 Phosphoprotein phosphatases reverse the actions of kinases and are independently regulated 625
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Principles of cell signaling - UT Southwestern

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