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Fundamental Technical Elements of Freeze-fracture/Freeze-etch in Biological Electron Microscopy
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The structural basis for membrane binding and pore formation by lymphocyte perforin.
Ruby H P Law1, Natalya Lukoyanova, Ilia Voskoboinik
1Department of Biochemistry and Molecular Biology, Monash University, Clayton, Melbourne, Victoria 3800, Australia.
Nature
|November 2, 2010
Summary
Natural killer cells use perforin to kill infected cells. Researchers determined the X-ray crystal structure of perforin, revealing its pore formation mechanism and unexpected MACPF domain orientation.
Area of Science:
- Immunology
- Structural Biology
- Cell Biology
Background:
- Natural killer cells and cytotoxic T lymphocytes eliminate virus-infected and cancerous cells.
- This process involves releasing perforin and granzymes to induce target cell death.
- Perforin mutations cause familial haemophagocytic lymphohistiocytosis (type 2 FHL), underscoring its critical role.
Purpose of the Study:
- To elucidate the mechanism of perforin pore formation.
- To determine the structural basis of perforin's function in cell-mediated cytotoxicity.
Main Methods:
- X-ray crystallography of monomeric murine perforin.
- Cryo-electron microscopy reconstruction of the perforin pore.
Main Results:
- The X-ray crystal structure revealed perforin's 'key-shaped' monomeric form with MACPF, EGF, and C2 domains.
- Cryo-electron microscopy showed perforin oligomerizes to form pores for granzyme delivery.
- Unexpectedly, the MACPF domain in the pore is oriented inside-out compared to related CDC proteins.
Conclusions:
- The study provides a detailed structural mechanism for perforin pore formation.
- Findings reveal the flexibility of the MACPF/CDC fold in immune defense.
- This work offers new insights into the assembly of pore-forming immune proteins like complement.
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