SH2 Domains Recognize Contextual Peptide Sequence Information to Determine Selectivity

Abstract

Selective ligand recognition by modular protein interaction domains is a primary determinant of specificity in signaling pathways. Src homology 2 (SH2) domains fulfill this capacity immediately downstream of tyrosine kinases, acting to recruit their host polypeptides to ligand proteins harboring phosphorylated tyrosine residues. The degree to which SH2 domains are selective and the mechanisms underlying selectivity are fundamental to understanding phosphotyrosine signaling networks. An examination of interactions between 50 SH2 domains and a set of 192 phosphotyrosine peptides corresponding to physiological motifs within FGF, insulin, and IGF-1 receptor pathways indicates that individual SH2 domains have distinct recognition properties and exhibit a remarkable degree of selectivity beyond that predicted by previously described binding motifs. The underlying basis for such selectivity is the ability of SH2 domains to recognize both permissive amino acid residues that enhance binding and non-permissive amino acid residues that oppose binding in the vicinity of the essential phosphotyrosine. Neighboring positions affect one another so local sequence context matters to SH2 domains. This complex linguistics allows SH2 domains to distinguish subtle differences in peptide ligands. This newly appreciated contextual dependence substantially increases the accessible information content embedded in the peptide ligands that can be effectively integrated to determine binding. This concept may serve more broadly as a paradigm for subtle recognition of physiological ligands by protein interaction domains. Molecular & Cellular Proteomics 9:2391-2404, 2010. The Src homology 2 (SH2)1 domain is the classic archetype for the large family of modular protein interaction domains that serve to organize a diverse array of cellular processes. As its name suggests, the SH2 domain was identified in the regulatory regions of non-receptor protein-tyrosine kinases (PTKs) of the Src family (1). The human genome encodes 110 SH2 domain proteins (2) that represent the primary mechanism for cellular signal transduction immediately downstream of PTKs. SH2 domains interact with phosphorylated tyrosinecontaining peptide sequences (3-6). In doing so, they couple activated PTKs to intracellular pathways that regulate many aspects of cellular communication in metazoans (7,8). In this manner, SH2 domains function alongside PTKs and proteintyrosine phosphatases to direct specificity in phosphotyrosine signaling. SH2 domain proteins play a critical role in development and have been linked to a wide range of human diseases including cancers, diabetes, and immunodeficiencies (2).Activation and recruitment of PTKs and protein-tyrosine phosphatases can control the spatial and temporal organization of phosphotyrosine signaling. However, signaling specificity in terms of downstream targets is entirely determined by phosphotyrosine-mediated recruitment events for which SH2 domains are responsible. Therefore, the binding selectivity of SH2 domains is critical for ...

Posttranslational protein knockdown coupled to receptor tyrosine kinase activation with phosphoPROTACs

“…Specifically, we tested whether a PROTAC can selectively distinguish between an activated versus a quiescent receptor tyrosine kinase-signaling pathway. Because phosphorylated tyrosine residues can serve as ligands for the binding of phosphotyrosine-binding (PTB) and Src homology 2 (SH2) domain-containing proteins (8), a cellpermeable PROTAC molecule that incorporates a specific tyrosine kinase substrate sequence may do the same (9). Thus, phosphorylation of such a PROTAC by a receptor tyrosine kinase (RTK) could generate a binding site for a downstream PTB or SH2 domain-containing protein and its subsequent recruitment to the ubiquitin/proteasome degradation pathway for destruction, thereby disrupting tyrosine kinase signaling ( Fig.…”

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“…Yaffe et al [22], in studying the phosphopeptide-binding protein 14-3-3, found that substrates adhered to one of two mutually exclusive sequence modes. Liu et al [23] have demonstrated that SH2 domains, which recognize and bind to phosphotyrosine-containing peptides, will vary in their specificity at some with a phosphotyrosine along with a leucine or a proline in the þ3 position. Proline at the þ2 position, however, is allowed only when the identity of the þ3 position is leucine and not proline.…”

Do low-affinity ErbB receptor protein interactions represent the base of a cell signaling iceberg?

“…The first assumption is that amino acids that contribute positively to an interaction are more important than amino acids that contribute negatively to binding. A recent study by Liu et al suggested that amino acids that block SH2 domain interaction may be as important in contributing to SH2 domain selectivity as amino acids that contribute to binding [18]. A second assumption is that the context of amino acids is not important in defining interaction specificity.…”