Mass spectrometry analysis of the oligomeric C1q protein reveals the B chain as the target of trypsin cleavage and interaction with fucoidan

C1q is a subunit of the C1 complex that triggers activation of the complement classical pathway through recognition and binding of immune complexes. C1q also binds to nonimmune ligands such as the sulfated polysaccharide fucoidan, a potent anticomplementary agent. C1q was submitted for the first time to mass spectrometry analysis, yielding insights into its assembly and its interaction with fucoidan. The MALDI-TOF mass spectrometry technique on membrane allowed partial preservation of noncovalent interactions, allowing precise analysis of its substructure and estimation of the C1q molecular weight at 459520-461883, with an average mass of 460793 g x mol(-1). The disulfide-linked A-B and C-C dimers as well as the noncovalent structural unit (A-B:C)-(C:B-A) were detected, providing experimental support to the C1q model based on covalent and noncovalent associations of six heterotrimers. Trypsin treatment of native C1q led to proteolysis of the B chain only, at a single cleavage site (Arg(109)) located in the globular region. Unlike DNA, fucoidan protected C1q from trypsin cleavage, indicating that this polysaccharide binds to the B moiety of the globular head. Given the involvement of the C1q globular heads in the recognition of IgG, this interaction may account for the observed anticomplementary activity of fucoidan.     

Interaction of fucoidan with the proteins of the complement classical pathway

Fucoidan inhibits complement by mechanisms that so far remain to be unraveled, and the objective of this work was to delineate the mode of inhibition by this sulfated polysaccharide. For that purpose, low molecular weight fractions of algal (Ascophyllum nodosum) fucoidan containing the disaccharide unit [-->3)-alpha-L-Fuc(2SO3(-))-(1-->4)-alpha-L-Fuc(2,3diSO3(-))-(1-->](n) have been studied. Gel co-affinity electrophoresis and a new affinity capillary electrophoresis (ACE) method have been implemented to characterize fucoidan-complement protein complexes. Fucoidan binds C1q, likely to its collagen-like region through interactions involving lysine residues, and then prevents the association of the C1r(2)-C1s(2) subunit, required to form the fully active C1. In addition to C1q, fucoidan forms a complex with the protein C4 as observed by ACE. The fucoidan inhibits the first steps of the classical pathway activation that is of relevance in view of the proinflammatory effects of the subsequent products of the cascade. This study shows that a high level of inhibitory activity can be achieved with low molecular weight carbohydrate molecules and that the potential applicability of fucoidan oligosaccharides for therapeutic complement inhibition is worthy of consideration.     

The effect of fucoidan on tyrosinase: computational molecular dynamics integrating inhibition kinetics

Fucoidan is a complex sulfated polysaccharide extracted from brown seaweed and has a wide variety of biological activities. In this study, we investigated the inhibitory effect of fucoidan on tyrosinase via a combination of inhibition kinetics and computational simulations. Fucoidan reversibly inhibited tyrosinase in a mixed-type manner. Time-interval kinetics showed that the inhibition was processed as first order with biphasic processes. For further insight, we simulated dockings with various sizes of molecular models (monomer to decamer) of fucoidan and showed that the best binding energy change results were obtained from the pentamer (?1.89?kcal/mol) and the hexamer (?1.97?kcal/mol) models of AutoDock Vina. The molecular dynamics simulation confirmed the binding mechanisms between tyrosinase and fucoidan and suggested that fucoidan mostly interacts with several residues including copper ions located in the active site. Our study suggests that fucoidan might be a potential natural antipigment agent.     

C1q binding to liposomes is surface charge dependent and is inhibited by peptides consisting of residues 14-26 of the human C1qA chain in a sequence independent manner

Complement activation by anionic liposomes proceeds by antibody-independent, C1q-initiated activation of the classical pathway. Purified C1q bound to anionic liposomes in an acidic lipid concentration-dependent manner. Saturation binding, but not the apparent association constant, was enhanced by increasing the cardiolipin content of the liposomes or decreasing either the pH or ionic strength of the reaction mixture. These observations indicate the involvement of electrostatic factors in the binding. A highly cationic region in the collagen-like domain of C1q comprised of residues 14-26 of the C1qA polypeptide chain was assessed for involvement in liposome binding. This region has previously been shown to mediate C1q binding to other immunoglobulin-independent activators of the classical pathway of complement. Peptides containing residues 14-26 of C1qA, denoted C1qA14-26, inhibited C1q binding to and complement activation by anionic liposomes. The inhibitory capacity of these cationic peptides had no sequence or conformation specificity. Rather, the amount of positive charge on the peptides was the determining factor. When present in excess, peptides with five cationic residues inhibited C1q binding and complement activation; however, C1q peptides with only two cationic residues did not. In addition to the C1qA14-26 region, other parts of C1q that contain cationic residues may also be involved in C1q binding to anionic liposomes.     

Chemical characterization and location of ionic interactions involved in the assembly of the C1 complex of human complement

The C1 complex of human complement comprises two loosely interacting subunits, C1q and the Ca²⁺-dependent C1s-C1r-C1r-C1s tetramer. With a view to gain information on the nature of the ionic interactions involved in C1 assembly, we have studied the effects of the chemical modifications of charged residues of C1q or the tetramer on their ability to reconstitute the C1 complex. Treatment of C1q with pyridoxal-5-phosphate, acetic anhydride, and citraconic anhydride, as well as with cyclohexanedione and diethylpyrocarbonate, inhibited its ability to associate with C1s-C1r-C1r-C1s. Treatment of the collagen-like fragments of C1q with the same reagents yielded the same effects. Treatment of C1s-C1r-C1r-C1s with 1-ethyl-3-[3-(dimethylamino) propyl] carbodiimide also prevented C1 assembly, through modification of acidic amino acids which were shown to be located in C1r. Further studies on the location of the interaction sites within C1q, using ligand-blotting and competition experiments with synthetic peptides, were unsuccessful, suggesting that these sites are contributed to by two or three of the C1q chains. It is concluded that C1 assembly involves interactions between acidic amino acids of C1r and lysine (hydroxylysine) and arginine residues located within the collagen-like region of C1q. Sequence comparison with mannan binding protein, another collagen-like molecule which binds the C1s-C1r-C1r-C1s tetramer, suggests Arg A38, and HyL B32, B65, and C29 of C1q as possible interaction sites.     

The effect of fucoidan on tyrosinase: computational molecular dynamics integrating inhibition kinetics

Fucoidan is a complex sulfated polysaccharide extracted from brown seaweed and has a wide variety of biological activities. In this study, we investigated the inhibitory effect of fucoidan on tyrosinase via a combination of inhibition kinetics and computational simulations. Fucoidan reversibly inhibited tyrosinase in a mixed-type manner. Time-interval kinetics showed that the inhibition was processed as first order with biphasic processes. For further insight, we simulated dockings with various sizes of molecular models (monomer to decamer) of fucoidan and showed that the best binding energy change results were obtained from the pentamer (-1.89?kcal/mol) and the hexamer (-1.97?kcal/mol) models of AutoDock Vina. The molecular dynamics simulation confirmed the binding mechanisms between tyrosinase and fucoidan and suggested that fucoidan mostly interacts with several residues including copper ions located in the active site. Our study suggests that fucoidan might be a potential natural antipigment agent.     

DNA binds and activates complement via residues 14-26 of the human C1q A chain.

The mechanism by which DNA activates the classical complement pathway was investigated, with emphasis upon the C1q binding sites involved. DNA bound to both the collagen-like and globular regions of C1q. Binding reactivity with DNA was retained after reduction/alkylation and sodium dodecyl sulfate treatment of C1q. DNA bound preferentially to the A chain of C1q. Binding sites for DNA were localized by using synthetic C1q A chain peptides to two cationic regions within residues 14-26 and 76-92, respectively. Peptides 14-26 and 76-92 avidly bound DNA in enzyme-linked immunosorbent and gel shift assays. Peptide 14-26 also precipitated with DNA and blocked its ability to bind C1q and activate C. Replacement of the two prolines with alanines or scrambling the order of the amino acids resulted in loss of ability of peptide 14-26 to inhibit C1q binding and complement activation by DNA; similar investigations showed a sequence specificity for peptide 76-92 as well. These experiments identify C1q A chain residues 14-26 as the major site, and residues 76-92 as a secondary site, through which DNA binds C1q and activates the classical complement pathway, and demonstrate that a peptide identical to residues 14-26 can modulate C1q binding and complement activation by DNA.     

The location of binding sites on C1q for DNA

Previous studies have suggested that C1q reacts with DNA via both the globular region of C1q (GR) and the collagen-like region of C1q (CLR). In this study, the binding of dsDNA and ssDNA to GR and CLR was quantitated by a solid-phase assay. Both dsDNA and ssDNA bound to the GR and CLR of C1q in an ionic strength-dependent manner. Under physiologic salt concentrations, however, dsDNA and ssDNA bound preferentially to CLR and not to GR. The binding of dsDNA to C1q was not affected by heat inactivation of C1q or its exposure to pH 4.45, which abolished the binding of heat-aggregated human IgG (AHG) with C1q. The preincubation of the solid-phase C1q with AHG did not decrease the binding of dsDNA or ssDNA to the solid-phase C1q. These results indicate that the major sites for binding DNA to C1q are located in the CLR of C1q and are not overlapping with those for AHG or immune complexes.     

Competitive inhibition of the classical complement pathway using exogenous single-chain C1q recognition proteins

Complement component C1q is a protein complex of the innate immune system with well-characterized binding partners that constitutes part of the classical complement pathway. In addition, C1q was recently described in the central nervous system as having a role in synapse elimination both in the healthy brain and in neurodegenerative diseases. However, the molecular mechanism of C1q-associated synapse phagocytosis is still unclear. Here, we designed monomer and multimer protein constructs, which comprised the globular interaction recognition parts of mouse C1q (globular part of C1q [gC1q]) as single-chain molecules (sc-gC1q proteins) lacking the collagen-like effector region. These molecules, which can competitively inhibit the function of C1q, were expressed in an Escherichia coli expression system, and their structure and capabilities to bind known complement pathway activators were validated by mass spectrometry, analytical size-exclusion chromatography, analytical ultracentrifugation, CD spectroscopy, and ELISA. We further characterized the interactions between these molecules and immunoglobulins and neuronal pentraxins using surface plasmon resonance spectroscopy. We demonstrated that sc-gC1qs potently inhibited the function of C1q. Furthermore, these sc-gC1qs competed with C1q in binding to the embryonal neuronal cell membrane. We conclude that the application of sc-gC1qs can reveal neuronal localization and functions of C1q in assays in vivo and might serve as a basis for engineering inhibitors for therapeutic purposes.     

Beta-amyloid fibrils activate the C1 complex of complement under physiological conditions: evidence for a binding site for A beta on the C1q globular regions.

Previous studies based on the use of serum as a source of C have shown that fibrils of beta-amyloid peptides that accumulate in the brain of patients with Alzheimer's disease have the ability to bind C1q and activate the classical C pathway. The objective of the present work was to test the ability of fibrils of peptide Abeta1-42 to trigger direct activation of the C1 complex and to carry out further investigations on the site(s) of C1q involved in the interaction with Abeta1-42. Using C1 reconstituted from purified C1q, C1r, and C1s, it was shown that Abeta1-42 fibrils trigger direct C1 activation both in the absence of C1 inhibitor and at C1 inhibitor:C1 ratios up to 8:0, i.e., under conditions consistent with the physiological context in serum. The truncated peptide Abeta12-42 and the double mutant (D7N, E11Q) of Abeta1-42 did not yield C1 activation, providing further evidence that the C1 binding site of beta-amyloid fibrils is located in the acidic N-terminal 1-11 region of the Abeta1-42 peptide. Binding studies performed using a solid phase assay provided strong evidence that C1q interacts with Abeta1-42 fibrils through its C-terminal globular regions. In contrast to previous studies based on a different experimental design, no significant involvement of the C1q collagen-like domain was detected. These findings were confirmed by additional experiments based on C1 activation and C4 consumption assays. These observations provide direct evidence of the ability of beta-amyloid fibrils to trigger activation of the classical C pathway and further support the hypothesis that C activation may be a component of the pathogenesis of Alzheimer's disease.     

Binding and complement activation by C-reactive protein via the collagen-like region of C1q and inhibition of these reactions by monoclonal antibodies to C-reactive protein and C1q

Ligand-complexed C-reactive protein (CRP), like aggregated or complexed IgG, can react with C1q and activate the classical C pathway. Whereas IgG is known to bind to the globular region and not to the collagen-like region (CLR) of C1q, the site of interaction of C1q with CRP has not been defined. CRP-trimers were prepared by cross-linking and found to bind to C1q and to activate the C system. Heat-aggregated IgG (Agg-IgG) did not block the binding of CRP-trimers to C1q, nor did CRP-trimers block binding of Agg-IgG to C1q, suggesting that CRP and IgG bind at different sites. ELISA and Western blot analysis showed that CRP-trimers bound to the CLR, whereas Agg-IgG bound only to the globular region; similarly, anti-CLR mAb inhibited binding of CRP-trimers to C1q whereas anti-globular region mAb did not. Reactivity with CRP-trimers as well as with Agg-IgG was retained after reduction/alkylation and SDS treatment of C1q. A group of 22 anti-CRP mAb directed against at least six distinct native-CRP epitopes and eight distinct neo-CRP epitopes was tested for ability to inhibit the CRP-CLR interaction; one mAb, anti-native CRP mAb 8D8, with strong inhibitory activity was identified. Fab' of 8D8 blocked binding of CRP-trimers to intact C1q as well as CLR, and also inhibited CRP (CRP-trimers and CRP-protamine complexes) induced C activation, but had no effect on C1q binding or C activation by Agg-IgG. These results indicate that a conformation-determined region on CRP binds to a sequence-determined region on the CLR of C1q in an interaction which leads to C activation. Anti-CRP and anti-C1q mAb that specifically inhibit this interaction are described.     

Role of Ca2+ in the electrostatic stability and the functional activity of the globular domain of human C1q

C1q is the recognition subunit of the classical pathway of the complement system and a major connecting link between classical pathway-driven innate immunity and IgG- or IgM-mediated acquired immunity. The basic structural subunit of C1q is composed of an N-terminal triple-helical collagen-like region and a C-terminal heterotrimeric globular head domain (gC1q) that is made up of individual A, B, and C chains. Recent crystallographic studies have revealed that the gC1q domain, which is the main target-binding region of C1q, has a compact and spherical heterotrimeric assembly, held together by both electrostatic and nonpolar interactions, with quasi-3-fold symmetry. A characteristic feature of the gC1q domain is the presence of a exposed Ca(2+) located near the apex. We have investigated, using theoretical and experimental approaches, the role of Ca(2+) in the electrostatic stability and target-binding properties of the native C1q as well as recombinant monomeric forms of the C-terminal regions of the A, B, and C chains. Here, we report that Ca(2+) primarily influences the target recognition properties of C1q toward IgG, IgM, C-reactive protein, and pentraxin 3. At pH 7.4, the loss of Ca(2+) leads to changes in the direction of electric moment from coaxial (where the putative C-reactive protein-binding site is located) to perpendicular to the molecular axis (toward the most likely IgG-binding site), which appears important for target recognition by C1q and subsequent complement activation.     

The inhibition of boar acrosin amidase activity by sulfated polysaccharides

Carbohydrate-protein interactions are known to be important in gamete interactions. We therefore investigated the inhibition of boar sperm acrosin amidase activity by carbohydrates. The sulfated polysaccharides fucoidan and dextran sulfate inhibited amide hydrolysis whereas dextran and various monosaccharides did not inhibit acrosin amidase activity. The kinetics of the inhibition corresponded to those characteristic when multiple forms of an enzyme are present. Such a kinetic result was consistent with the presence of the known autolytically produced forms of acrosin. It was previously shown that sulfated polysaccharides inhibit sperm-egg binding and that acrosin binds carbohydrate. We propose that the sulfated polysaccharide inhibition of acrosin amidase activity observed here is causally related to the previously observed sulfated polysaccharide inhibition of sperm binding to the zona pellucida.     

Role of complement component C1q in the IgG-independent opsonophagocytosis of group B streptococcus.

We investigated the role of complement component C1q in the IgG-independent opsonophagocytosis of type III group B Streptococcus (GBS) by peripheral blood leukocytes. We report that C1q binds to type III GBS both in normal human serum deficient in IgG specific for type III capsular polysaccharide and in a low-ionic strength buffer. The dissociation constant Kd ranged from 2.0 to 5.5 nM, and the number of binding sites Bmax ranged from 630 to 1360 molecules of C1q per bacterium (CFU). An acapsular mutant strain of GBS bound C1q even better than the wild type, indicating that the polysaccharide capsule is not the receptor for C1q. In serum, binding of C1q to GBS was associated with activation of the classical complement pathway. However, normal human serum retained significant opsonic activity after complete depletion of C1q, suggesting that the serum contains a molecule that is able to replace C1q in opsonization and/or complement activation. Mannan-binding lectin, known to share some functions with C1q, appeared not to be involved, since its depletion from serum had little effect on opsonic activity. Excess soluble C1q or its collagen-like fragment inhibited phagocytosis mediated by normal human serum, suggesting that C1q may compete with other opsonins for binding to receptor(s) on phagocytes. We conclude that, although C1q binds directly to GBS, C1q binding is neither necessary nor sufficient for IgG-independent opsonophagocytosis. The results raise the possibility that additional unknown serum factor(s) may contribute to opsonization of GBS directly or via a novel mechanism of complement activation.     

Antibody-independent interaction between the first component of human complement, C1, and the outer membrane of Escherichia coli D31 m4.

The heptoseless mutant of Escherichia coli, E. coli D31 m4, binds C1q and C1 at 0 degrees C and at low ionic strength (I0.07). Under these conditions, the maximum C1q binding averages 3.0 X 10(5) molecules per bacterium, with a Ka of 1.4 X 10(8) M-1. Binding involves the collagen-like region of C1q, as shown by the capacity of C1q pepsin-digest fragments to bind to E. coli D31 m4, and to compete with native C1q. Proenzyme and activated forms of C1 subcomponents C1r and C1s and their Ca2+-dependent association (C1r-C1s)2 do not bind to E. coli D31 m4. In contrast, the C1 complex binds very effectively, with an average fixation of 3.5 X 10(5) molecules per bacterium, and a Ka of 0.25 X 10(8) M-1, both comparable with the values obtained for C1q binding. C1 bound to E. coli D31 m4 undergoes rapid activation at 0 degrees C. The activation process is not affected by C1-inhibitor, and only slightly inhibited by p-nitrophenyl p'-guanidinobenzoate. No turnover of the (C1r-C1s)2 subunit is observed. Once activated, C1 is only partially dissociated by C1-inhibitor. Our observations are in favour of a strong association between C1 and the outer membrane of E. coli D31 m4, involving mainly the collagen-like moiety of C1.     

Structure and anti-dengue virus activity of sulfated polysaccharide from a marine alga

A sulfated polysaccharide, named fucoidan, from the marine alga Cladosiphon okamuranus is comprised of carbohydrate units containing glucuronic acid and sulfated fucose residues. Here we found this compound potently inhibits dengue virus type 2 (DEN2) infection. Viral infection was inhibited when DEN2, but not other serotypes, was pretreated with fucoidan. A carboxy-reduced fucoidan derivative in which glucuronic acid was converted to glucose did not inhibit viral infection. Elimination of the sulfated function group from fucoidan significantly attenuated the inhibitory activity on DEN2 infection with <1% fucoidan. DEN2 particles bound exclusively to fucoidan, indicating that fucoidan interacts directly with envelope glycoprotein (EGP) on DEN2. Structure-based analysis suggested that Arg323 of DEN2 EGP, which is conformationally proximal to one of the putative heparin binding residues, Lys310, is critical for the interaction with fucoidan. In conclusion, both the sulfated group and glucuronic acid of fucoidan account for the inhibition of DEN2 infection.     

Electrospray ionization mass spectrometry of oligosaccharides derived from fucoidan of Ascophyllum nodosum

Algal fucoidan is a complex sulfated polysaccharide whose structural characterization requires powerful spectroscopic methodologies. While most of the structural investigations reported so far have been performed using NMR as the main spectroscopic method, we report herein data obtained by negative electrospray ionization mass spectrometry. MS analysis has been carried out on oligosaccharides obtained by partial hydrolysis of fucoidan from the brown algae Ascophyllum nodosum. Oligosaccharide mixtures were fractionated by size exclusion chromatography, which allowed the analysis of oligomers ranging from monosaccharide to pentasaccharide. Monosaccharides were detected as monosulfated as well as disulfated forms. Besides, part of the oligosaccharides exhibited a high content of sulfate, evidencing that fucoidan contains disulfated fucosyl units. Fragmentation experiments yielded characteristic fragment ions indicating that the fucose units are mainly 2-O-sulfated. This study demonstrates that highly sulfated oligosaccharides from fucoidan can be analyzed by ESIMS which gives additional information about the structure of this highly complex polysaccharide.     

Monthly changes in the content and monosaccharide composition of fucoidan from <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> (Laminariales,Phaeophyta)

Crude fucoidan was extracted from the brown alga Undaria pinnatifida collected monthly from April to last July in Peter the Great Bay (Japan Sea, Russia). The amount of crude fucoidan rose markedly from April to June–July (from 3.2 to 16.0% dry weight) as the plant matures. An analysis of the monosaccharide composition of the fucoidan extracted showed that the alga synthesized polysaccharides with various structures which were dependent on the algae age. In juvenile plants collected in April–May, this was represented by sulfated manno-galactofucan containing up to 19–28 mol% of mannose and about 20 mol% of galactose, whereas in matured plants collected in June–July, the polysaccharide was represented by a sulfated galactofucan containing more than 38 mol% galactose. It is postulated that the production of sori causes a subsequent effect on fucoidan synthesis and leads to an enhanced of crude fucoidan content and an increased molar concentration of galactose. Crude fucoidan content in sporophylls increased 5 times, and galactose content in this polysaccharide rose s1.6 times with sori formation. The structural characteristics of the fucoidan extracted from sporophylls of Undaria collected in July were also studied. The fractionation of crude fucoidan on DEAE-Sephadex A-25 gave two fractions, F1 and F2 in equal quantities. F1 was characterized as manno-galactofucan sulfate and F2 was galactofucan sulfate. The molecular weights of both fractions were in a range of 30–80 kDa. Analysis of fucoidan structure using ESI-FTICR mass spectrometry showed the presence of mixed oligosaccharides consisting of fucose and galactose. Presumably, the polysaccharide molecules contain blocks built up of successively linked residues of fucose and galactose. These blocks are built from two to five or more residues of monosaccharides. According to IR-spectroscopy data, the main portion of sulfates is located at C2; in addition, sulfate esters are also present at C4 on the fucose and C3 and C6 of the galactose units.     

Fibronectin binding to complement subcomponent C1q. Localization of their respective binding sites.

The interaction of purified human plasma fibronectin with the C1q subcomponent of complement was investigated by using a solid-phase radiobinding assay. 125I-fibronectin binding to native C1q, purified collagen domain (C1q-c) or globular domain (C1q-g) was compared. When the purified domains were insolubilized by binding to plastic, the C1q-c exhibited 59% of the binding demonstrated with intact C1q, whereas the C1q-g exhibited 35% of the binding. N-Terminal sequencing of the globular domain showed that a sequence of seven collagen-like amino acids was retained on each chain of the C1q-g fragment. 125I-fibronectin binding to C1q could be inhibited equally well by fluid-phase C1q and C1q-c, but not by fluid-phase C1q-g, implying that the collagen-like region retained on the C1q-g is masked in the fluid phase. In addition, studies were performed to determine which subunit(s) of C1q bind(s) fibronectin. The percentages of fibronectin bound by the A, B, and C chain of C1q were found to be 38, 21 and 41% respectively. Inhibition studies with purified 200-180 kDa, 50 kDa or 29 kDa fragments of fibronectin show that the binding site on fibronectin for C1q is the 50 kDa gelatin-binding domain.     

Evidence that complement protein C1q interacts with C-reactive protein through its globular head region

C1q acts as the recognition unit of the first complement component, C1, and binds to immunoglobulins IgG and IgM, as well as to non-Ig ligands, such as C-reactive protein (CRP). IgG and IgM are recognized via the globular head regions of C1q (C1qGR), whereas CRP has been postulated to interact with the collagen-like region (C1qCLR). In the present study, we used a series of nine mAbs to C1q, five directed against C1qGR and four against C1qCLR, to inhibit the interaction of C1q with CRP. The F(ab')(2) of each of the five mAbs directed against C1qGR inhibited binding of C1q to polymerized IgG. These five mAbs also successfully inhibited the interaction of C1q with CRP. Moreover, these five mAbs inhibited C1 activation by CRP as well as by polymerized IgG in vitro. In contrast, none of the four mAbs against C1qCLR inhibited C1q interaction with CRP or IgG, or could reduce activation of complement by CRP or polymerized IgG. These results provide the first evidence that the interaction of C1q with CRP or IgG involves sites located in the C1qGR, whereas sites in the CLR do not seem to be involved in the physiological interaction of C1q with CRP.