Diamine-induced dissociation of the first component of human complement, C1

Abstract

Lysine has been shown to inhibit spontaneous and antibody-dependent C1 activation. This paper demonstrates that lysine does not prevent autoactivation of purified Clr. 20 mM lysine, 1,2-diaminoethane, 1,3-diaminopropane, 1,Cdiaminobutane or 1,5-diaminopentane are able to dissociate C1 into its two entities, C l q and the calciumdependent Clr,-Cls, complex. Ig-ovalbumin insoluble complexes bearing C1 are also dissociated by lysine and the above-mentioned diamines used at the same concentration: Clq remains bound to the immune complexes whereas the Clr,-Cls, complex is partially solubilized.The effect of lysine or diamines is not due to a competition with calcium for calcium-binding sites, as increasing concentrations of calcium even slightly increase the dissociation due to the amines.The dissociative effect is dependent on the carbon chain length of the diamines, with an optimum for 1,3-diaminopropane. It is also dependent on the relative 'cis-position' of the amino groups in the diamines. Polyamines such as spermine and spermidine are also able to dissociate C1 with even a higher efficiency than lysine and putrescine.Thus, a diamine-induced 'structural inhibition' of C1 is demonstrated, of potential interest for a pharmacological control of complement activation.The complement system contributes to the non-specific immune defence of the organism, as it can be triggered by different stimuli such as immune complexes, polyanions, bacterial or viral membranes, exogenous proteases [l 1. Activation of the system can proceed by two different pathways, the classical and the alternative pathway, and consists of a cascade of proteolytic reactions. Biological effects such as cytolysis or inflammation are the most evident expression of the activation.Activation of the classical pathway implies, as a primary event, the activation of C1, a calcium-dependent complex of three different proteins, Clq, C l r and Cls. The structure of C1 is now relatively well known and appears to consist of two entities: a tight Clr,-Cls2 complex, in loose association with Clq, under physiological conditions [2 -71. The activation of C1 proceeds through a sequential activation of C l r and CIS in which the initial autoactivation of C l r appears the most meaningful event. The autocatalytic potential is expressed in purified C l r itself, which is able to autoactivate upon incubation at 37 "C, in the presence of EDTA [8 -lo]. Inside C1, this autocatalytic potential is modulated by the association with the two other subcomponents: Cls and Clq, as the activation energy for C1 activation [Ill is lower than the activationenergy for C l r activation [8]. Incontrast, in the ClrNomenclature and Abbreviations. The nomenclature of complement components is that recommended by the World Health Organization [20] ; activated components are indicated by an overbar. SDS, sodium dodecyl sulphate; cadaverine, 1 $diaminopentane ; putrescine, 1 .Cdiaminobutane ; spermidine, W(3-aminopropyl)-1,4-diaminobutane ; spermine, N,N'-bis(3 aminopropy1)-I ,4-diaminobutane; a...

Domain structure and associated functions of subcomponents C1r and C1s of the first component of human complement.

“…4) for the calcium-dependent dimerization of Cls, which occurs in the absence of Clr (6,7,11,17). It (4,25). In agreement with comparative neutron diffraction studies on Clq and C1 (26), this led us to the conclusion that in the Cl complex the (Cir, Cls)2 subunit adopts a nonlinear conformation corresponding to its activatable state.…”

“…Clq is considered as the recognition subunit and its structure has been elucidated by chemical and electron microscopic studies (1,2). (Cir, Cls)2, the catalytic subunit, is released from C1 by treatment with citrate (3) or diamino compounds (4) or at high ionic strength (5). Subcomponents Cir and Cls are both serine proteases that, at the monomeric level, appear very similar: the zymogen forms are both single-chain proteins of molecular weights ranging from 83,000 to 95,000, converted on activation into two disulfide-linked polypeptide chains, A and B, of respective molecular weights 58,000 and 28,000-35,000 (6)(7)(8)(9)(10)(11).…”

Structure and Activation of C1: Current Concepts

“…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. Recently, dissociation of Cl by diam ines has been demonstrated [14], in the pres ence of 20mM lysine, 1,2-diaminoethane, 1,3-diaminopropane, 1,4-diaminobutane or 1,5-diaminopentane ( fig. 1).…”

Polyamines Can Increase Resistance ofNeisseria gonorrhoeaeto Mediators of the Innate Human Host Defense

“…We do not yet know if early steps in CCP activation or steps in MAC formation are impaired by spermine, but we suggest that spermine masks sites on the bacterial surface which influence either one or both processes. Since previous work documented the ability of spermine to dissociate the C1 complex (38), it is likely that the capacity of 8 mM spermine to significantly reduce the bactericidal effect of NHS and to abolish C8 binding to whole gonococci can be explained by this mechanism. Clearly, however, the results from our hemolytic assays using antibody-sensitized sheep blood erythrocytes indicate that residual CCP activity remains in such serum but that the level is insufficient to kill bacteria.…”

“…The capacity of spermine to inhibit NHS-mediated activity against gonococci and E. coli suggested that its presence in the SBA influenced CCP activity. Indeed, earlier studies that used 20 mM or greater concentrations of spermine showed that this polyamine can inhibit the formation of the C1 complex and prevent activation of the downstream CCP cascade (10,20,38). Using a modified protocol of the EZ complement CH50 test (Diamedix, Miami, FL), we found that 8 mM spermine, but not 1 mM spermine, significantly decreased the hemolytic activity of 1% (vol/vol) but not 10% (vol/vol) NHS against antibodysensitized sheep erythrocytes (Fig.…”