Background
Obesity affects over 650 million adults worldwide and is a leading cause of cardiovascular disease, diabetes, and cancer. GPR75, an orphan G protein-coupled receptor (GPCR), has been genetically validated as an anti-obesity target — loss-of-function carriers have significantly lower BMI and reduced risk of obesity-related diseases.
The Challenge
At only 56 kDa (receptor + nanobody), the GPR75 complex was near the lower size limit for cryo-EM single particle analysis. Key challenges included:
- Small size: Weak signal-to-noise ratio in micrographs
- Conformational flexibility: GPCRs are inherently dynamic
- Sample preparation: Achieving homogeneous active-state complex
- Preferred orientation: Membrane proteins often adopt limited views on grids
Shuimu's Approach
- Nanobody stabilization: A conformation-specific nanobody was developed to lock GPR75 in its active signaling state
- GraFuture™ graphene oxide grids: Used to overcome preferred orientation by providing a hydrophilic support
- Optimized data collection: 300kV Titan Krios G3i with BioQuantum GIF and K3 camera, collecting >5000 movies
- Advanced processing: Iterative 2D/3D classification with heterogeneous refinement
- First-ever structure of GPR75 in active conformation
- Revealed the orthosteric ligand binding pocket architecture
- Identified key residues for G protein coupling specificity
- Provided a structural template for virtual screening and rational drug design
Results
Impact
This structure enables pharmaceutical companies to design selective GPR75 agonists that could become a new class of anti-obesity drugs. The work demonstrated that cryo-EM SPA can tackle GPCR complexes previously considered too small.
Publication
Cryo-EM complex structure of active GPR75 with a nanobody — bioRxiv, 2022. Read the paper →