Structural insight into allosteric modulation of protease-activated receptor 2

Area of Science:

• Biochemistry
• Molecular Biology
• Pharmacology

Background:

• Protease-activated receptors (PARs) are GPCRs crucial in inflammatory responses and disease pathogenesis.
• Developing effective PAR2 antagonists has been challenging due to difficulties in targeting the receptor effectively.

Purpose of the Study:

• To elucidate the structural basis of PAR2 antagonism by determining crystal structures of PAR2 in complex with antagonists and an antibody.
• To provide a foundation for the rational design of novel PAR2-targeted therapeutics.

Main Methods:

• X-ray crystallography was employed to determine the structures of PAR2 bound to two distinct antagonists (AZ8838, AZ3451) and a blocking antibody fragment.
• Functional and binding assays were conducted to characterize antagonist kinetics and mechanisms of action.

Main Results:

• The antagonist AZ8838 was observed to bind within an occluded pocket near the extracellular surface, exhibiting slow binding kinetics.
• The antagonist AZ3451 was found to bind at a remote allosteric site, suggesting a mechanism of preventing necessary receptor rearrangements.
• A blocking antibody fragment was shown to bind the extracellular surface, sterically hindering tethered ligand access.

Conclusions:

• The determined structures offer detailed insights into distinct modes of PAR2 antagonism.
• These findings provide a structural framework for developing selective PAR2 antagonists for therapeutic applications in various diseases.