Cryo-EM structure of an activated GPR4-Gs signaling complex

Area of Science:

• Structural Biology
• Biochemistry
• Molecular Biology

Background:

• G protein-coupled receptor 4 (GPR4) is a proton-sensing receptor involved in physiological responses.
• Overactivation of GPR4 in acidic tumor microenvironments and inflammation highlights its pathological relevance.
• The 3D structure and detailed proton-sensing mechanisms of GPR4 remain largely unelucidated.

Purpose of the Study:

• To determine the cryo-electron microscopy (cryo-EM) structures of active zebrafish GPR4 at different pH values.
• To investigate the role of extracellular ionizable residues in GPR4's proton-sensing capacity.
• To elucidate the structural basis for signal propagation in GPR4.

Main Methods:

• Cryo-electron microscopy (cryo-EM) for structural determination.
• Cell-based assays to assess proton-sensing capacity.
• Analysis of conserved residues within proton-sensing GPCRs (psGPCRs).

Main Results:

• Reported cryo-EM structures of active zebrafish GPR4 at pH 6.5 and 8.5.
• Identified extracellular histidine and acidic residues influencing proton sensing, though less significantly than transmembrane triad residues.
• Revealed a cluster of aromatic residues in the orthosteric pocket potentially mediating signal propagation.

Conclusions:

• The study provides high-resolution structures of GPR4, offering a structural framework for understanding psGPCR function.
• Extracellular ionizable residues contribute moderately to GPR4's pH sensitivity.
• Aromatic residue interactions within the orthosteric pocket are implicated in GPR4 signal transduction.