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Modulation of the SH2 binding specificity and kinase activity of Src by tyrosine phosphorylation within its SH2
D R Stover1, P Furet, N B Lydon
1Pharmaceutical Research Division, Ciba-Geigy Limited, Basel, Switzerland.
Abstract:
The Src family of kinases are held in an inactive state by interaction of their SH2 domain with a C-terminal phosphotyrosine. Dephosphorylation of this site can reactivate Src; however, recent evidence suggests that activation can also occur without dephosphorylation. In this study, platelet-derived growth factor receptor phosphorylation of Src on Tyr-213 specifically blocked binding of its SH2 domain to a phosphopeptide corresponding to the C-terminal regulatory sequence, while binding to other sequences, such as the platelet-derived growth factor receptor or a peptide from the epidermal growth factor receptor, was unaffected. Consequently, Src was activated over 50-fold. This is the first demonstration of regulation of a SH2 domain specificity by post-translational modification and is likely to be a general mechanism for regulation of all Src-like kinases.
Insights
Src kinases are regulated by phosphorylation. Platelet-derived growth factor receptor (PDGFR) phosphorylation of Src at Tyr-213 alters SH2 domain binding, activating Src over 50-fold.
Area of Science:
- Molecular Biology
- Cell Signaling
- Kinase Regulation
Background:
- Src family kinases are crucial signaling enzymes.
- These kinases are typically maintained in an inactive state via intramolecular interaction.
- Reactivation can occur through dephosphorylation or other mechanisms.
Purpose of the Study:
- To investigate the mechanism of Src kinase activation.
- To explore the role of post-translational modification in regulating SH2 domain specificity.
- To determine if phosphorylation can alter Src's interaction with its regulatory phosphotyrosine site.
Main Methods:
- Utilized phosphopeptide binding assays.
- Investigated the effect of platelet-derived growth factor receptor (PDGFR) phosphorylation on Src.
- Measured Src activation in response to specific phosphorylation events.
Main Results:
- PDGFR phosphorylation of Src on Tyr-213 specifically disrupted SH2 domain binding to the C-terminal regulatory phosphopeptide.
- Binding to other peptides, including PDGFR and epidermal growth factor receptor (EGFR) peptides, remained unaffected.
- Src kinase activity was increased over 50-fold due to this modification.
Conclusions:
- This study demonstrates a novel mechanism for Src kinase regulation.
- Post-translational modification of Src can directly alter SH2 domain specificity.
- This phosphorylation-dependent regulation of SH2 domain binding is a likely general mechanism for Src-like kinases.