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Activation of the Src-family tyrosine kinase Hck by SH3 domain displacement
I Moarefi1, M LaFevre-Bernt, F Sicheri
1Laboratories of Molecular Biophysics, Howard Hughes Medical Institute, The Rockefeller University, New York 10021, USA.
Insights
HIV-1 Nef protein binding to the Hck SH3 domain significantly increases tyrosine kinase activity. This interaction reveals a novel regulatory mechanism for Hck, a key enzyme in immune cells.
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Hck is a Src family tyrosine kinase crucial in hematopoietic cells.
- Src kinases are regulated by C-terminal tyrosine phosphorylation, involving SH2 domains.
- The SH3 domain also influences Hck activity through intramolecular interactions.
Purpose of the Study:
- To compare the roles of SH2 and SH3 domains in modulating Hck activity.
- To investigate the effect of HIV-1 Nef protein on Hck regulation.
Main Methods:
- Structural analysis of Hck's downregulated form.
- Investigated C-terminally phosphorylated Hck activity.
- Assessed the impact of HIV-1 Nef protein binding to Hck SH3 domain.
Main Results:
- HIV-1 Nef protein binds to the Hck SH3 domain with high affinity.
- Nef binding to Hck SH3 domain causes a significant increase in catalytic activity.
- SH3 domain engagement by Nef is more potent in activating Hck than SH2 domain interaction.
Conclusions:
- The Hck SH3 domain plays a critical role in enzyme activation.
- HIV-1 Nef protein binding to Hck SH3 domain represents a novel activation mechanism.
- This study elucidates a new pathway for tyrosine kinase regulation.
Abstract:
The protein Hck is a member of the Src family of non-receptor tyrosine kinases which is preferentially expressed in haematopoietic cells of the myeloid and B-lymphoid lineages. Src kinases are inhibited by tyrosine-phosphorylation at a carboxy-terminal site. The SH2 domains of these enzymes play an essential role in this regulation by binding to the tyrosine-phosphorylated tail. The crystal structure of the downregulated form of Hck has been determined and reveals that the SH2 domain regulates enzymatic activity indirectly; intramolecular interactions between the SH3 and catalytic domains appear to stabilize an inactive form of the kinase. Here we compare the roles of the SH2 and SH3 domains in modulating the activity of Hck in an investigation of the C-terminally phosphorylated form of the enzyme. We show that addition of the HIV-1 Nef protein, which is a high-affinity ligand for the Hck SH3 domain, to either the downregulated or activated form of Hck causes a large increase in Hck catalytic activity. The intact SH3-binding motif in Nef is crucial for Hck activation. Our results indicate that binding of the Hck SH3 domain by Nef causes a more marked activation of the enzyme than does binding of the SH2 domain, suggesting a new mechanism for regulation of the activity of tyrosine kinases.