Conformational characterization of the intrinsically disordered protein Chibby: Interplay between structural elements...

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• Chibby (Cby) protein antagonizes Wnt signaling by inhibiting β-catenin/Tcf-Lef interactions.
• Full characterization of Cby's C-terminal coiled-coil domain was limited by NMR line broadening.

Purpose of the Study:

• To structurally and functionally characterize the C-terminal half of the Chibby (Cby) protein.
• To investigate the role of Cby's structural elements in protein stability and interaction with TC-1.

Main Methods:

• Hydrogen/deuterium exchange-mass spectrometry (HDX-MS) to analyze Cby's C-terminal half.
• Circular Dichroism (CD) and Nuclear Magnetic Resonance (NMR) spectroscopy.
• Isothermal Titration Calorimetry (ITC) for binding affinity measurements.

Main Results:

• Cby comprises a disordered N-terminal half, a coiled-coil domain, and a C-terminal extension.
• Coiled-coil structure is maintained upon deletion of disordered regions.
• Cby's C-terminal half alone exhibits higher affinity for TC-1 than full-length Cby.

Conclusions:

• Cby's disordered regions modulate the binding affinity of its C-terminal coiled-coil domain to TC-1.
• HDX-MS provides valuable insights into the structure of partially disordered proteins like Cby.