SH2 domain Structure

Due to the ready accessibility of SH2 domains by molecular biology techniques, numerous experimentally determined 3D structures of SH2 domains derived by X-ray crystallography as well as heteronuclear multidimensional NMR spectroscopy are known today. The current version of the protein structure database, accessible to the scientific community by, e.g., the Internet (http //www.rcsb.org/pdb/) contains around 80 entries of SH2 domain structures and complexes thereof. Today, the SH2 domain structures of Hck [62], Src [63-66], Abl [67], Grb2 [68-71], Syp [72], PLCy [73], Fyn [74], SAP [75], Lck [76,77], the C- and N-terminal SH2 domain ofp85a [78-80], and of the tandem SH2 domains Syk [81,82], ZAP70 [83,84], and SHP-2 [85] are determined. All SH2 domains display a conserved 3D structure as can be expected from multiple sequence alignments (Fig. 4). The common structural fold consists of a central three-stranded antiparallel ft sheet that is occasionally extended by one to three additional short strands (Fig. 5). This central ft sheet forms the spine of the domain which is flanked on both sides by regular a helices [49, 50,60]. 

The first SH2 domain structures to be solved were those of the Src tyrosine kinase (Waksman et al., 1992), the Abl tyrosine kinase (Overduin et al., 1992), and the N-terminal SH2 domain of the p85 subunit of the PI 3 -kinase (N-p85 SH2 domain) (Booker et al., 1992). These structures revealed the architecture of SH2 domains. The SH2 domain fold is relatively simple it consists of a central antiparallel (3-sheet flanked by... 

The structure of the N-SHP-2 SH2 domain in complex with peptides based on the platelet-derived growth factor receptor (PDGFR) and the adapter protein IRSl revealed that the interaction between the protein and the peptide is extended to the residue 5 positions C-terminal to the pTyr (Lee et al., 1994). In these structures, the +3 position binding pocket is opened in order to create such an extended interface. The interactions between the peptide and the protein are primarily hydro-phobic. For example, a Phe at the +5 position of the IRSl peptide binds between the EF and BG loops and interacts with the He at the +3 position (Lee et al., 1994). Other than this extended interface, the interactions in this structure are reminiscent of those observed in the Src SH2 domain structure the peptide binds in an extended conformation perpendicular to the central (3-sheet. 

Figure 13.30 Ribbon diagram of the structure of Src tyrosine kinase. The structure is divided in three units starting from the N-terminus an SH3 domain (green), an SH2 domain (blue), and a tyrosine kinase (orange) that is divided into two domains and has the same fold as the cyclin dependent kinase described in Chapter 6 (see Figure 6.16a). The linker region (red) between SH2 and the kinase is bound to SH3 in a polyproline helical conformation. A tyrosine residue in the carboxy tail of the kinase is phosphorylated and bound to SH2 in its phosphotyrosine-binding site. A disordered part of the activation segment in the kinase is dashed. (Adapted from W. Xu et al.. Nature 385 595-602, 1997.)...

Src tyrosine kinase contains both an SH2 and an SH3 domain linked to a tyrosine kinase unit with a structure similar to other protein kinases. The phosphorylated form of the kinase is inactivated by binding of a phosphoty-rosine in the C-terminal tail to its own SH2 domain. In addition the linker region between the SH2 domain and the kinase is bound in a polyproline II conformation to the SH3 domain. These interactions lock regions of the active site into a nonproductive conformation. Dephosphorylation or mutation of the C-terminal tyrosine abolishes this autoinactivation. 

Waksman, G., et al. Binding of a high affinity phosphoty-rosyl peptide to the Src SH2 domain crystal structures of the complexed and peptide-free forms. Cell 72 779-790, 1993. 

Insulin Receptor. Figure 1 Structure and function of the insulin receptor. Binding of insulin to the a-subunits (yellow) leads to activation of the intracellular tyrosine kinase ((3-subunit) by autophosphorylation. The insulin receptor substrates (IRS) bind via a phospho-tyrosine binding domain to phosphorylated tyrosine residues in the juxtamembrane domain of the (3-subunit. The receptor tyrosine kinase then phosphorylates specific tyrosine motifs (YMxM) within the IRS. These tyrosine phosphorylated motifs serve as docking sites for some adaptor proteins with SRC homology 2 (SH2) domains like the regulatory subunit of PI 3-kinase. 

Molecular insight into the protein conformation states of Src kinase has been revealed in a series of x-ray crystal structures of the Src SH3-SH2-kinase domain that depict Src in its inactive conformation [7]. This form maintains a closed structure, in which the tyrosine-phosphorylated (Tyr527) C-terminal tail is bound to the SH2 domain (Fig. 2). The x-ray data also reveal binding of the SH3 domain to the SH2-kinase linker [adopts a polyproline type II (PP II) helical conformation], providing additional intramolecular interactions to stabilize the inactive conformation. Collectively, these interactions cause structural changes within the catalytic domain of the protein to compromise access of substrates to the catalytic site and its associated activity. Significantly, these x-ray structures provided the first direct evidence that the SH2 domain plays a key role in the self-regulation of Src. 

Figure 2 Depiction of the active ( open ) and inactive ( closed ) conformations of Src kinase based on the analysis of x-ray structures of c-Src tyrosine kinase crystallized in its inactive state [7]. The stabilization of the inactive conformation is influenced by multiple events including intramolecular binding of the tyrosine-phosphorylated C-terminus tail to the SH2 domain as well as interactions between the SH3 domain and the SH2-kinase linker. CT, C-terminal NT, N-terminal. 

Plummer MS, Holland DR, Shahripour A, Lunney EA, Fergus JH, Marks JS, McConnell P, Mueller WT, Sawyer TK. Design, synthesis, and cocrystal structure of a nonpeptide Src SH2 domain ligand. J Med Chem 1997 40 3719-3725. 

Lunney EA, Para KS, Rubin JR, Humblet C, Fergus JH, Marks JS, Sawyer TK. Structure-based design of a novel series of nonpeptide ligands that bind to the pp60c"src SH2 domain. J Am Chem Soc 1997 119 12471-12476. 

Metcal CA III, Eyermann CJ, Bohacek RS, Haraldson C, Varkhedkar VM, Lynch B, Bartlett C, Violette S, Sawyer TK. Structure-based design and solid-phase parallel synthesis of phosphorylated nonpeptides to explore hydro-phobic binding at the Src SH2 domain. J Comb Chem 2000 2 305-313. 

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