Fluid phase activation of proenzymic C1r purified by affinity chromatography

Villiers, Christian; Duplaa, A.M.; Arlaud, Gérard J.; Colomb, Maurice G.

doi:10.1016/0167-4838(82)90299-0

Cited by 30 publications

(25 citation statements)

References 29 publications

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“…Dissociation of the two subunits of Cl can also be achieved by citrate or diamino compounds. As reported by Villiers et al [13], a Clr2-Cls2 complex can be bound from diluted citrated plasma to a Sepharose-IgG anti-Cls column. In these conditions, the calcium ions present in the Clr2-Cls2 complex appear inaccessible to chelators such as EDTA, and Clr is removed from the adsorbent only by lowering the pH to 5.0.…”

Section: Evidence For Two Distinct Entities In CLmentioning

confidence: 88%

Structure and Activation of C1: Current Concepts

Mg¹,

Gj²,

Cl³

1984

Complement

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Section: Evidence For Two Distinct Entities In CLmentioning

confidence: 88%

Structure and Activation of C1: Current Concepts

Mg¹,

Gj²,

Cl³

1984

Complement

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“…A comparable inhibitory effect of Ca2+ was observed when incubation was performed at 30 'C; after 1 h at this temperature, a CIr preparation undergoing 420% activation the presence of EDTA yielded only 7 % activation in the presence of Ca2+. It has been shown that autoactivation of isolated Cl r involves two distinct mechanisms: an intramolecular activation reaction catalysed by the proenzyme, and an intermolecular reaction catalysed by activated Cir generated in the course of the reaction (Villiers et al, 1982a). With a view to evaluating the susceptibility of each mechanism to Ca2+ ions, the effect of Ca2+ was compared with that produced by iPr2P-F, a serine-proteinase inhibitor known to react covalently with the active-site serine residue of Cir (Villiers et al, 1982a), and therefore expected to prevent the intermolecular activation reaction.…”

Section: Ca2+ Slows Down Clr Autoactivation Through Inhibition Of Thementioning

confidence: 99%

Activation of human complement serine-proteinase C1r is down-regulated by a Ca2+-dependent intramolecular control that is released in the C1 complex through a signal transmitted by C1q

Thielens

Illy

Bally

et al. 1994

Biochemical Journal

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The activation of human C1, a Ca(2+)-dependent complex proteinase comprising a non-enzymic protein, C1q, and two serine proteinases, C1r and C1s, is based primarily on the intrinsic property of C1r to autoactivate. The aim of the present study was to investigate the mechanisms involved in the regulation of C1r autoactivation, with particular attention to the role of Ca2+ ions. Spontaneous activation of proenzyme C1r was observed upon incubation in the presence of EDTA, whereas Ca2+ ions reduced markedly the activation process. Several lines of evidence indicated that Ca2+ inhibited the intramolecular activation reaction but had little or no effect on the intermolecular activation reaction. C1q caused partial release of this inhibitory effect of Ca2+. Complete stabilization of C1r in its proenzyme form was obtained upon incorporation within the Ca(2+)-dependent C1s-C1r-C1r-C1s tetramer, and a comparable effect was observed when C1s was replaced by its Ca(2+)-binding alpha-fragment. Both tetramers, C1s-C1r-C1r-C1s and C1s alpha-C1r-C1r-C1s alpha, readily associated with C1q to form 16.0 S and 14.7 S complexes respectively in which C1r fully recovered its activation potential. Both complexes showed indistinguishable activation kinetics, indicating that the gamma B catalytic region of C1s plays no role in the mechanism that triggers C1r activation in C1. The collagen-like fragments of C1q retained the ability to bind to C1s-C1r-C1r-C1s, but, in contrast with intact C1q, failed to induce C1r activation in the resulting complex at temperatures above 25 degrees C. On the basis of these observations it is proposed that activation of the serine-proteinase domain of C1r is controlled by a Ca(2+)-dependent intramolecular mechanism involving the Ca(2+)-binding alpha-region, and that this control is released in C1 by a signal originating in C1q and transmitted through the C1q/C1r interface.

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“…(1987). Translation was for 2 h at 30 "C. Immunoprecipitation of the translated proteins was carried out, according to Reboul et al (1986), with anti-CIs antibodies (Villiers et al, 1982) or commercially available antibodies (anti-lysozyme; Calbiochem). SDS/polyacrylamide (12.5%, w/v) gel electrophoresis was performed under reducing conditions.…”

Section: Dna Probesmentioning

confidence: 99%

Identification of a human non-interferon lymphokine activating monocyte complement biosynthesis

Drouet

Reboul

Colomb

1989

Biochemical Journal

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A monocyte-stimulating activity produced by mitogen-induced mononuclear cells has been defined by its ability to enhance the synthesis in vitro of complement C1 subcomponents, C2 and C3. A lymphokine responsible for this activity was purified from culture supernatants of peripheral blood mononuclear cells activated by staphylococcal enterotoxin A. From 0.5 litre of supernatant the purification procedure [(NH4)2SO4 precipitation, phenyl-Sepharose chromatography and preparative electrofocusing] yielded about 100 pmol of purified lymphokine. Its pI is 7.9 and its Mr, estimated by SDS/polyacrylamide-gel electrophoresis, is 14,600, 27,000 and 56,000, the high-Mr species representing oligomeric forms of the Mr-14,600 molecule. Its amino acid analysis reveals a high percentage of hydrophobic amino acids (34%); the absence of histidine residues suggests that it is a novel monocyte-activating lymphokine. It enhances C1r and C1s biosynthesis at a pretranslational level. From its structure and activity this lymphokine appears different from gamma-interferon.

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Fluid phase activation of proenzymic C1r purified by affinity chromatography

Cited by 30 publications

References 29 publications

Structure and Activation of C1: Current Concepts

Structure and Activation of C1: Current Concepts

Activation of human complement serine-proteinase C1r is down-regulated by a Ca2+-dependent intramolecular control that is released in the C1 complex through a signal transmitted by C1q

Identification of a human non-interferon lymphokine activating monocyte complement biosynthesis

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