C1-inhibitor prevents PEG fractionation-induced, EDTA-resistant activation of mouse complement.
Fractionation of mouse serum by precipitation with a critical amount of polyethylene glycol 6000 (PEG; 11% w/v) results in a classical and alternative pathway-independent activation of the terminal complement route. The activation can take place after the separation of an activating principle together with the terminal route components from a natural regulator. The isolation and identification of the regulatory component preventing this activation in serum, is subject of this paper. The regulator was purified by fractionated PEG-precipitation (15-25%), followed by heparin-Sepharose affinity, Mono Q anion-exchange, and Superose 12 gel filtration chromatography. The regulator appeared to be a single-chain protein with a Mr of 96 k. A protein with similar activity purified from human serum had a Mr of 104 k and was functionally and antigenically indistinguishable from C1-INH. The mouse 96 k protein inhibited C1-esterase activity indicating that this protein is indeed C1-INH. Mouse C1-INH regulates the PEG fractionation-induced bypass activation of complement, but does not interfere with the assembly or the lytic activity of membrane attack complexes. alpha 2-Macroglobulin appeared also to be capable of inhibiting the PEG-precipitation-induced activation process, but with lower efficiency.
Eijkman-Winkler Laboratory of Medical Microbiology, Department of Experimental Microbiology, Faculty of Medicine, University of Utrecht, The Netherlands.
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van den Berg
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Aerts
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van Dijk
D000818:Animals; D000990:Antithrombin III / Q000494:pharmacology; D002845:Chromatography; D003167:Complement Activation / Q000187:drug effects*; D003174:Complement C1 Inactivator Proteins / Q000502:physiology*; D003173:Complement C1s / Q000276:immunology / Q000302:isolation & purification; D015941:Complement Hemolytic Activity Assay; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005779:Immunodiffusion; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D011092:Polyethylene Glycols; D053491:Proteinase Inhibitory Proteins, Secretory; D011506:Proteins; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D000515:alpha 1-Antitrypsin / Q000494:pharmacology; D000511:alpha-Macroglobulins / Q000494:pharmacology
D003174:Complement C1 Inactivator Proteins; D053491:Proteinase Inhibitory Proteins, Secretory; D011506:Proteins; D015842:Serine Proteinase Inhibitors; D000515:alpha 1-Antitrypsin; D000511:alpha-Macroglobulins; D011092:Polyethylene Glycols; D000990:Antithrombin III; D003173:Complement C1s