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Updated: Mar 31, 2026
Optimized Negative Staining: a High-throughput Protocol for Examining Small and Asymmetric Protein Structure by Electron Microscopy
Published on: August 15, 2014
Structure of Full-Length Human PDGFRβ Bound to Its Activating Ligand PDGF-B as Determined by Negative-Stain Electron
Po-Han Chen1, Vinzenz Unger2, Xiaolin He1
1Department of Biochemistry and Molecular Genetics, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA.
Abstract:
Members of the receptor tyrosine kinases (RTKs) regulate important cellular functions such as cell growth and migration, which are key steps in angiogenesis, in organ morphogenesis and in the unregulated states, cancer formation. One long-standing puzzle regarding RTKs centers on how the extracellular domain (ECD), which detects and binds to growth factors, is coupled with the intracellular domain kinase activation. While extensive structural works on the soluble portions of RTKs have provided critical insights into RTK structures and functions, lack of a full-length receptor structure has hindered a comprehensive overview of RTK activation. In this study, we successfully purified and determined a 27-Å-resolution structure of PDGFRβ [a full-length human platelet-derived growth factor receptor], in complex with its ligand PDGF-B. In the ligand-stimulated complex, two PDGFRβs assemble into a dimer via an extensive interface essentially running along the full-length of the receptor, suggesting that the membrane-proximal region, the transmembrane helix and the kinase domain of PDGFRβ are involved in dimerization. Major structural differences are seen between the full-length and soluble ECD structures, rationalizing previous experimental data on how membrane-proximal domains modulate receptor ligand-binding affinity and dimerization efficiency. Also, in contrast to the 2-fold symmetry of the ECD, the intracellular kinase domains adopt an asymmetric dimer arrangement, in agreement with prior observations for the closely related KIT receptor. In essence, the structure provides a first glimpse into how platelet-derived growth factor receptor ECD, upon ligand stimulation, is coupled to its intracellular domain kinase activation.
Insights
This study reveals the full-length structure of platelet-derived growth factor receptor beta (PDGFRβ) bound to its ligand. The structure elucidates how the extracellular domain couples with intracellular kinase activation through receptor dimerization.
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- Receptor tyrosine kinases (RTKs) regulate crucial cellular processes including growth, migration, and angiogenesis.
- A key challenge in RTK research is understanding the coupling mechanism between the extracellular ligand-binding domain and intracellular kinase activation.
- Previous studies lacked a full-length RTK structure, limiting a comprehensive view of receptor activation.
Purpose of the Study:
- To determine the structure of full-length human platelet-derived growth factor receptor beta (PDGFRβ) in complex with its ligand PDGF-B.
- To elucidate the structural basis of PDGFRβ activation and dimerization upon ligand binding.
- To provide insights into the coupling mechanism between the extracellular and intracellular domains of RTKs.
Main Methods:
- Purification of full-length human PDGFRβ.
- Complex formation with its ligand PDGF-B.
- Determination of the 27-Å-resolution structure of the PDGFRβ-PDGF-B complex using cryo-electron microscopy.
Main Results:
- A 27-Å-resolution structure of full-length PDGFRβ complexed with PDGF-B was determined.
- Ligand-stimulated PDGFRβ forms an extensive dimer interface along the full receptor length, involving membrane-proximal regions, transmembrane helix, and kinase domain.
- Significant structural differences exist between full-length and soluble extracellular domains, explaining modulation of ligand-binding and dimerization.
- Intracellular kinase domains exhibit an asymmetric dimer arrangement, contrasting with the extracellular domain's 2-fold symmetry.
Conclusions:
- The determined structure offers the first detailed view of full-length PDGFRβ activation.
- It reveals how ligand binding induces dimerization and couples the extracellular domain to intracellular kinase activation.
- The findings rationalize previous experimental data and provide a structural basis for understanding RTK signaling.
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