Headless C1q: a new molecular tool to decipher its collagen-like functions

Area of Science:

• Immunology
• Structural Biology
• Biochemistry

Background:

• Complement component C1q initiates the classical complement pathway.
• C1q comprises collagen-like (CLR) and globular (GR) regions, crucial for target recognition and immune response.
• The GRs bind diverse targets, while CLRs interact with cellular/soluble partners.

Purpose of the Study:

• To generate the first recombinant human C1q lacking its globular heads.
• To substitute the C1q GR with the noncollagenous domain 2 (nc2) of type IX collagen.
• To assess the assembly, folding, and functional properties of the resulting CLR_nc2 recombinant protein.

Main Methods:

• Protein engineering: Substitution of C1q GR with type IX collagen nc2 domain.
• Expression in EXPI293 mammalian cells.
• Characterization using mass spectrometry, mass photometry, and electron microscopy.
• Interaction analysis via surface plasmon resonance with C1r/C1s and MAp44.

Main Results:

• Successfully generated and produced the CLR_nc2 recombinant protein.
• Confirmed correct assembly and folding of CLR_nc2.
• Demonstrated that CLR_nc2 retains the functional properties of the native C1q CLR in binding known ligands.

Conclusions:

• The CLR_nc2 recombinant protein serves as a valuable tool for studying C1q's collagen-like region functions.
• This engineered protein maintains the essential interaction capabilities of the native CLR.
• Provides insights into the structure-function relationship of C1q in complement activation.