The functional properties of the secreted form of C4 (C4(s)), which has a M(r) ~5000 greater than the predominant C4 molecule found in plasma (C4(p)), two incompletely processed two-chain C4 molecules (β - α + γ and β + α - γ), and the extracellular C4 precursor (designated pro-C4(E)) were evaluated. All four molecules are secreted in parallel by a human hepatoma-derived cell line (Hep G2). Secretion of hemolytically active C4 is linear up to ~12 hr, peaks at 24 hr, and then progressively decreases over the next 48 hr. This loss of C4(s) functional activity parallels the proteolytic conversion of C4(s) to C4b(s). To compare the hemolytic efficiencies of C4(s) and C4(p), a solid-phase competitive radioimmunoassay was developed to permit measurement of the small quantities of C4 antigen in these cultures. The hemolytic efficiencies of C4(s) and C4(p) were similar. These results indicate that extracellular processing of C4(s) to C4(p) does not modulate the hemolytic activity of the molecule. Consistent with their ability to bind methylamine, both the α(s)-chain and the α - γ subunit undergo denaturation-induced autolysis. The extracellular and intracellular pro-C4 molecules are also sensitive to autolytic cleavage. Interestingly, the β - α subunit is resistant to autolysis. In experiments in which C4(s) and C4(p) were cleaved by C1sβ to C4b(s), C4(β - α + γ), C4(β + α - γ), and pro-C4(E) were resistant to C1s̄ cleavage and thus hemolytically inactive relative to C4(s). These data indicate that processing of C4 to a three-chain structure is required to provide the proper conformation for efficient activation by C1.

Original languageEnglish
Pages (from-to)1967-1971
Number of pages5
JournalJournal of Immunology
Issue number4
StatePublished - 1984


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