The opsonic fragment of the third component of human complement (C3)

Area of Science:

• Immunology
• Cell Biology
• Biochemistry

Background:

• Phagocytosis of bacteria by immune cells is a critical defense mechanism.
• Complement system activation, particularly involving C3, plays a vital role in opsonization, enhancing phagocytosis.
• The precise nature of the C3 fragment responsible for opsonization of lipopolysaccharide (LPS)-coated particles was not fully characterized.

Purpose of the Study:

• To investigate the role of complement C3 in the phagocytosis of LPS-coated particles by human peripheral blood phagocytes.
• To identify and characterize the specific C3 fragment involved in the opsonization process.
• To elucidate the mechanism by which C3 facilitates particle recognition and ingestion.

Main Methods:

• Opsonization of paraffin oil droplets coated with E. coli LPS using fresh human serum.
• Assay of particle ingestibility by phagocytes using spectrophotometry.
• Radioactive labeling of C3 with [125I] and analysis of bound radioactivity.
• Treatment of opsonized particles with various agents (temperature, pH, detergents, enzymes, antibodies) to assess C3 fragment stability and function.
• Electrophoretic and tryptic peptide autoradiographic analysis of C3-derived polypeptides.

Main Results:

• Phagocytosis of LPS-coated particles required C3 deposition (opsonization) mediated by the properdin system.
• The rate of C3 binding correlated directly with particle ingestibility.
• Opsonized particles retained ingestibility and bound [125I]C3 after exposure to harsh conditions, indicating a stable C3 fragment.
• Specific treatments (trypsin, SDS, anti-C3 F(ab')2) diminished ingestibility and/or removed radioactivity.
• Analysis revealed the opsonic C3 fragment had a molecular weight of ~140,000 Da, composed of disulfide-linked 70,000 Da subunits, and was a smaller derivative of C3, distinct from C3b.

Conclusions:

• Complement C3 opsonization, mediated by the properdin pathway, is essential for phagocyte ingestion of LPS-coated particles.
• The opsonic fragment is a stable, smaller derivative of C3, not C3b, highlighting a specific role in phagocytic recognition.
• Understanding this C3 fragment's structure and function provides insights into complement-mediated immunity and phagocytosis mechanisms.