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Regulatory system of guinea-pig complement C3b: two factor I-cofactor proteins on guinea-pig peritoneal granulocytes
Insights
Researchers identified two distinct complement factor I cofactor activities in guinea pig granulocytes. These species-specific factors cleave complement C3b, potentially identifying membrane cofactor protein and C3b/C4b receptor roles.
Area of Science:
- Immunology
- Protease biochemistry
Background:
- Complement factor I is crucial for regulating the complement system by inactivating C3b.
- Identifying cofactors for factor I is essential for understanding complement regulation.
Purpose of the Study:
- To identify and characterize factor I cofactor activities in guinea pig granulocytes.
- To investigate the species specificity of these cofactor activities.
Main Methods:
- Solubilization of guinea pig peritoneal granulocytes.
- Chromatofocusing to separate cofactor activities.
- Proteolytic cleavage assays using fluorescently labeled guinea pig C3.
- SDS-PAGE to analyze protein components.
Main Results:
- Two distinct factor I cofactor activities were isolated, eluting at pH 7.6-7.1 (neutral) and pH 5.7 (acidic).
- Both fractions cleaved guinea pig C3b but not human C3b, indicating species specificity.
- The neutral fraction contained 55 kDa and 42 kDa proteins, while the acidic fraction had a 160 kDa protein.
Conclusions:
- Guinea pig granulocytes possess distinct factor I cofactor activities.
- These cofactors are likely involved in complement regulation and may represent guinea pig membrane cofactor protein and C3b/C4b receptor.
Abstract:
Complement factor I is a plasma protease serving for proteolytic inactivation of C3b together with its cofactor. We have identified two factor I-cofactor activities in solubilized extracts of guinea-pig peritoneal granulocytes using guinea-pig factor I (Igp) and fluorescent-labeled methylamine-treated guinea-pig C3 (f-C3(MA)gp). One of these eluted from a chromatofocusing column between pH 7.6-7.1, and the other at about pH 5.7. These two cofactor fractions both interacted with Igp and, to a lesser degree, with human factor I (Ihu) on C3(MA)gp cleaving it into an inactive C3bi analogue, but did not cleave methylamine-treated human C3 (C3(MA)hu) together with Igp or Ihu. These factors are therefore species specific. The neutral and acidic fractions with cofactor activity contained C3(MA)gp-binding proteins with a doublet of 55 kDa and 42 kDa, and a singlet of 160 kDa, respectively, on SDS-PAGE. These proteins may be membrane cofactor protein (MCP) and C3b/C4b receptor (CR1) of guinea-pigs.