Structural Basis for Ca2+-mediated Interaction of the Perforin C2 Domain with Lipid Membranes

Area of Science:

• Immunology
• Molecular Biology
• Structural Biology

Background:

• Natural killer cells and cytotoxic T-lymphocytes use perforin and granzymes to eliminate infected cells.
• Perforin forms pores in target cell membranes for granzyme delivery, a process initiated by its C2 domain's calcium-dependent membrane interaction.
• The molecular mechanisms underlying perforin's C2 domain interaction with membranes remain incompletely understood.

Purpose of the Study:

• To elucidate the molecular mechanisms of calcium (Ca2+) and lipid binding to perforin's C2 domain.
• To investigate the structural rearrangements induced by Ca2+ binding in the C2 domain.

Main Methods:

• Nuclear Magnetic Resonance (NMR) spectroscopy
• X-ray crystallography
• Calcium titrations and dodecylphosphocholine micelle experiments

Main Results:

• Multiple Ca2+ ions bind to perforin's calcium-binding regions, activating it for membrane interaction.
• The affinities of several Ca2+ binding sites were determined.
• Ca2+ binding induces significant structural rearrangements in the C2 domain's calcium-binding region 1 (CBR1).

Conclusions:

• Ca2+ binding, particularly at the weakest affinity site, triggers conformational changes in the C2 domain.
• These Ca2+-induced structural changes facilitate perforin's subsequent interaction with lipid membranes.
• This study provides key molecular insights into the calcium-dependent activation of perforin for cell-mediated cytotoxicity.