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Covalent linkage of C3 to properdin during complement activation
L Y Whiteman1, D B Purkall, S Ruddy
1Department of Microbiology and Immunology, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298, USA.
European Journal of Immunology
|May 1, 1995
Summary
The alternative complement pathway forms a novel 160 kDa complex of properdin (P) and C3. This complex, involving a covalent linkage, is crucial for complement activation and impacts hemolytic activity.
Area of Science:
- Immunology
- Biochemistry
Background:
- The alternative complement pathway is a critical part of the innate immune system.
- Properdin (P) and C3 are key proteins involved in this pathway's activation.
Purpose of the Study:
- To investigate the formation and nature of complexes between properdin (P) and C3 during alternative complement pathway activation.
- To characterize the functional significance of the P-C3 complex.
Main Methods:
- Enzyme-linked immunosorbent assay (ELISA) to detect P-C3 complexes.
- Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting with biotinylated properdin and C3 antibodies.
- Radiolabeling (125I) of P and C3 in purified reagent mixtures followed by SDS-PAGE and autoradiography.
- Hydroxylamine treatment to assess covalent linkages.
Main Results:
- A 160 kDa complex containing both properdin (P) and C3 was identified during alternative pathway activation.
- This complex was shown to involve a covalent linkage between P and the C3 alpha chain.
- The formation of the P-C3 complex was confirmed in both activated serum and purified protein systems.
- The purified complex reduced factor B hemolytic activity.
Conclusions:
- Properdin (P) covalently binds to C3 via its alpha chain during alternative complement pathway activation, forming a 160 kDa complex.
- This novel P-C3 complex plays a role in the regulation of complement activation and hemolytic activity.