# The structural basis for PAR1 biased signaling

> **NIH NIH R21** · CASE WESTERN RESERVE UNIVERSITY · 2021 · $201,250

## Abstract

PROJECT SUMMARY/ABSTRACT
G-protein coupled receptors (GPCRs) elicit complex downstream signaling cascades by activating Gαq, Gα12/13,
Gαi, or arrestin. There is a growing appreciation that biased agonists can dictate which signaling pathways are
activated downstream of the receptor. Protease activated receptors (PARs) are the primary means by which
proteases initiate intracellular signaling. PARs are activated by cleavage of the N-terminus to generate a tethered
ligand. PAR1 has unique cleavage sites for multiple proteases that can lead to a panel of unique tethered ligands.
These ligands are endogenous biased agonists that trigger specific signaling pathways. The molecular basis for
this is not known. The long-term goals of this research program are to define how PARs mediate context specific
signaling in endothelial cells, platelets and other cells. This project seeks to develop a foundation for
understanding the molecular basis for PAR activation mechanisms that govern normal physiological responses.
The overall objective of this proposal is to 1.) define the tethered ligand binding site(s) for PAR1 activated by
three endogenous activators thrombin, APC, and MMP1 2.) uncover the structural basis for PAR1 biased
signaling 3.) determine how these sites cooperate to mediate specific physiological signaling events. Our overall
hypothesis is that PAR1 adopts specific conformations due to distinct ligand binding sites for each of the tethered
ligands dictating which signaling pathways are activated. The scientific premise is based on the recent success
of our experimental design that incorporates amide hydrogen/deuterium (H/D) exchange with purified PARs to
determine how the tethered ligand influences the overall conformation. Molecular modeling will independently
determine the ligand binding site(s). Finally, identified regions will be tested in cell signaling assays using a panel
of PAR1 mutants to verify the importance on cell signaling. Our innovative approach will identify the previously
unknown endogenous ligand binding sites for each of the tethered ligands generated by thrombin, APC, and
MMP1. At the completion of these studies will define the potential conformations of PAR1 with endogenous
activators. These experiments will provide the first structural insights as to how a single receptor can have
opposite signaling outcomes under normal physiological conditions.

## Key facts

- **NIH application ID:** 10241452
- **Project number:** 5R21HL154026-02
- **Recipient organization:** CASE WESTERN RESERVE UNIVERSITY
- **Principal Investigator:** Marvin Thomas Nieman
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $201,250
- **Award type:** 5
- **Project period:** 2020-09-01 → 2023-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10241452

## Citation

> US National Institutes of Health, RePORTER application 10241452, The structural basis for PAR1 biased signaling (5R21HL154026-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10241452. Licensed CC0.

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