Question

Many proteins contain phosphotyrosine-binding SH2, SH3 and PTB domains, yet the interaction of these proteins with appropriate taget molecules is exquisitely specific. What is the basis of such specificity?

Answer 1

An SH2 or PTB domain typically recognizes a pTyr residue within the context of a specific amino acid sequence. This characteristic allows for the distinct binding of SH2- and PTB-containing proteins to activated RTKs and to other tyrosine phosphorylated signaling effectors. These domains are found in proteins of diverse function, and on RTK activation their recruitment to pTyr sites results in the activation of their host proteins, and the stimulation of downstream signaling pathways. As noted, these proteins usually contain other catalytic and noncatalytic domains that establish their functions, and can be used to categorize them into distinct groups.

Adaptor proteins and docking/scaffolding proteins lack intrinsic catalytic activity, but act to assemble signaling complexes capable of selectively stimulating downstream pathways. Adaptors containing both SH2 and SH3 domains use the SH3 domain(s) to aggregate signaling effectors, whereas the SH2 domain mediates recruitment to the active RTK and assembly of the signaling complex. For example, the SH3 domains of Nck bind to effectors involved in the organization of the cytoskeleton (such as N-WASP and PAK) to link the active RTK to the cytoskeleton. Scaffolding proteins recruit signaling effectors through linear motifs, including pTyr-containing binding sites, and localized assembly of the signaling complex is directed by the scaffold’s PTB or SH2 domain. Shc and IRS-1, for instance, localize to pTyr motifs in an active RTK via their PTB domains and bind the Grb2/Sos complex following phosphorylation of internal YXN motifs, allowing for subsequent activation of the Ras-MAPK pathway.

In combination with a catalytic domain, the SH2 domains of kinases, phosphatases, guanine nucleotide exchange factors (GEFs), GTPase activating proteins (GAPs), and the phospholipase PLC-γ generally recruit these proteins to their substrates. Such a substrate can be either a tyrosine-phosphorylated protein or a distinct substrate juxtaposed to a tyrosine-phosphorylated protein. Within an enzyme, an SH2 domain can also allow for intramolecular binding and dynamic regulation of enzymatic activity, and this concept is discussed later in this article. In addition, enzymes containing domains capable of binding secondary messengers (such as PH domains binding phosphatidylinositol 3,4,5-triphosphate (PIP3) or C1 domains binding diacylglycerol (DAG)/phorbol esters) tend to show a higher level of specificity in both localization and activation.

Finally, in the STAT family of transcription factors, SH2 domains enable recruitment to active RTKs; on subsequent RTK-mediated phosphorylation, the SH2 domain mediates STAT dimerization and activation . Thus, specific binding of SH2 and PTB domains plays a major role in the localized assembly and activation of signaling effectors. In the sections that follow, we describe in detail how the specific localization of a protein is achieved through the various modes of SH2 binding.