Structural and functional properties of Grb2 SH2 dimer in CD28 binding.

Area of Science:

• Molecular Biology
• Structural Biology
• Immunology

Background:

• Growth factor receptor-bound protein 2 (Grb2) is a key adaptor protein in cellular signal transduction.
• The Src homology 2 (SH2) domain of Grb2 binds to phosphorylated CD28, a critical step for T cell costimulatory signaling.

Purpose of the Study:

• To investigate the structural and functional properties of Grb2 SH2 domain dimer and monomer forms.
• To understand the role of Grb2 SH2 dimer formation in CD28 binding and T cell signaling.

Main Methods:

• Purification of Grb2 SH2 dimer and monomer.
• Size exclusion chromatography and thermal stability analysis (circular dichroism).
• CD28 phosphopeptide binding assays and crystal structure analysis of Grb2 SH2.

Main Results:

• Both Grb2 SH2 dimer and monomer exist as stable states, with the dimer dissociating around 50°C.
• The Grb2 SH2 dimer showed a threefold higher affinity for CD28 phosphopeptide compared to the monomer, likely due to avidity.
• Crystal structures revealed monomeric and dimeric forms of Grb2 SH2, with dimer formation involving C-terminal helix swapping mediated by Trp121.

Conclusions:

• Stable Grb2 SH2 dimer formation via C-terminal helix swapping enhances CD28 binding affinity.
• The Trp121 residue is critical for Grb2 SH2 dimer formation and subsequent CD28 binding.
• These findings provide structural insights into Grb2-mediated T cell costimulatory signaling.