Human adenosine A2A receptor binds calmodulin with high affinity in a calcium-dependent manner

Area of Science:

• Biochemistry
• Structural Biology
• Molecular Pharmacology

Background:

• G-protein-coupled receptors (GPCRs) mediate cellular responses to external signals.
• The adenosine A2A receptor (A2AR) has a unique, long, and largely uncharacterized intracellular C-terminus.
• Understanding GPCR C-terminus structure and interactions is vital for signal transduction research.

Purpose of the Study:

• To elucidate the structure and dynamics of the A2AR C-terminus.
• To characterize the interaction between the A2AR C-terminus and calmodulin.
• To investigate the functional implications of this complex in receptor signaling.

Main Methods:

• Native gel electrophoresis and analytical gel filtration.
• Isothermal titration calorimetry (ITC).
• NMR spectroscopy and small-angle X-ray scattering (SAXS).

Main Results:

• The A2AR C-terminus is intrinsically disordered and flexible.
• High-affinity binding (Kd = 98 nM) of the C-terminus to calmodulin was observed without significant domain conformational changes.
• Calmodulin binds to helix 8 of A2AR in a calcium-dependent manner, influencing A2AR's association with lipid vesicles.

Conclusions:

• The A2AR C-terminus is a dynamic region that interacts with calmodulin.
• This interaction is calcium-dependent and modulates A2AR membrane association.
• Putative calmodulin-binding sites were identified in other GPCRs, suggesting a broader regulatory mechanism.