Single-molecule kinetics of pore assembly by the membrane attack complex.

Area of Science:

• Immunology
• Molecular Biology
• Biophysics

Background:

• The membrane attack complex (MAC) is crucial for pathogen lysis via complement system activation.
• The sequential assembly of MAC proteins (C5b-C9) leads to pore formation, but rate-limiting steps remain unclear.

Purpose of the Study:

• To elucidate the kinetic steps governing MAC assembly using advanced imaging techniques.
• To understand the role of individual protein incorporation in MAC pore formation.

Main Methods:

• Utilized rapid atomic force microscopy (AFM) for real-time imaging of MAC assembly on lipid bilayers.
• Investigated protein-lipid interactions and oligomerization dynamics during MAC formation.

Main Results:

• MAC proteins oligomerize within the membrane, differing from bacterial pore-forming toxins.
• C5b-7 complex interacts with the lipid bilayer before C8 recruitment.
• The incorporation of the first C9 molecule represents the kinetic bottleneck in MAC assembly.
• Subsequent rapid C9 polymerization efficiently completes the pore structure.

Conclusions:

• Identified the rate-limiting step in MAC formation as the initial C9 incorporation.
• Established the kinetic basis for MAC assembly, highlighting C9 as the key regulatory component.
• Provided mechanistic insight into CD59's protective role by defining the temporal window for halting MAC assembly at C9 insertion.