Identity of the Segment of Human Complement C8 Recognized by Complement Regulatory Protein CD59

Lockert, Dara H.; Kaufman, Kenneth M.; Chang, Chi-Pei; Hüsler, Thomas; Sodetz, James M.; Sims, P.

doi:10.1074/jbc.270.34.19723

Cited by 57 publications

(46 citation statements)

References 21 publications

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“…MAC formation itself is regulated through interaction with CD59, a widely distributed, membrane-anchored glycoprotein that binds to C8␣ and C9 and thereby restricts MAC assembly and function. Earlier studies showed the CD59-binding site in C8␣ is within a segment defined by residues 334 -385 in TMH2 (52). More recently, it was suggested the binding site lies within residues 350 -355 (53).…”

Section: Discussionmentioning

confidence: 99%

Structure of Human C8 Protein Provides Mechanistic Insight into Membrane Pore Formation by Complement

Lovelace

Cooper

Sodetz

et al. 2011

Journal of Biological Chemistry

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C8 is one of five complement proteins that assemble on bacterial membranes to form the lethal pore-like "membrane attack complex" (MAC) of complement. The MAC consists of one C5b, C6, C7, and C8 and 12-18 molecules of C9. C8 is composed of three genetically distinct subunits, C8␣, C8␤, and C8␥. The C6, C7, C8␣, C8␤, and C9 proteins are homologous and together comprise the MAC family of proteins. All contain N-and C-terminal modules and a central 40-kDa membrane attack complex perforin (MACPF) domain that has a key role in forming the MAC pore. Here, we report the 2.5 Å resolution crystal structure of human C8 purified from blood. This is the first structure of a MAC family member and of a human MACPF-containing protein. The structure shows the modules in C8␣ and C8␤ are located on the periphery of C8 and not likely to interact with the target membrane. The C8␥ subunit, a member of the lipocalin family of proteins that bind and transport small lipophilic molecules, shows no occupancy of its putative ligand-binding site. C8␣ and C8␤ are related by a rotation of ϳ22°with only a small translational component along the rotation axis. Evolutionary arguments suggest the geometry of binding between these two subunits is similar to the arrangement of C9 molecules within the MAC pore. This leads to a model of the MAC that explains how C8-C9 and C9-C9 interactions could facilitate refolding and insertion of putative MACPF transmembrane ␤-hairpins to form a circular pore. Assembly of the "membrane attack complex" (MAC)3 of complement on the surface of Gram-negative bacteria and other pathogenic organisms involves the sequential interaction of complement proteins C5b, C6, C7, C8, and C9 (1-3). Association of the first four components produces a membranebound tetrameric C5b-8 complex, which then initiates the recruitment and sequential binding of 12-18 C9 molecules to form a cylindrical transmembrane pore (supplemental Fig. S1). Pore formation leads to loss of membrane integrity and lysis of the cell under attack.The sequence of interactions leading to MAC formation is well defined; however, the mechanism by which the MAC disrupts membrane organization is poorly understood. C6, C7, the C8␣ and C8␤ subunits, and C9 are homologous and together comprise the "MAC family" of proteins (4, 5). Until now, structures have not been determined for any of these proteins. All contain N-and C-terminal modules and a central 40-kDa "membrane attack complex/perforin" (MACPF) domain. The MACPF domain was named as such because of sequence similarity between the MAC family proteins and perforin. The modules range in number from three to eight; all are small domains of 40 -60 amino acids that contain multiple disulfide bonds (supplemental Fig. S2). One type of module, thrombospondin type 1 (TSP1), contains several mannosylated tryptophans (6).Several hundred MACPF-containing proteins have been identified; however, functions are known for only a few. MACPF proteins exhibit limited sequence similarity, but all contain the MACPF signature motif ((Y/...

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Section: Discussionmentioning

confidence: 99%

Structure of Human C8 Protein Provides Mechanistic Insight into Membrane Pore Formation by Complement

Lovelace

Cooper

Sodetz

et al. 2011

Journal of Biological Chemistry

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“…Human CD59 is an 18-kDa glycosylphosphatidylinositollinked membrane glycoprotein, widely expressed in all circulating cells and most tissues, that serves as an inhibitor of the C5b-9 MAC of human complement (9,17,39,48). The complement inhibitory activity of CD59 depends on its ability to bind to C8 in the C5b-8 complex (30) and to C9 between Cys359 and Cys384 (21), thereby inhibiting ion channel formation by C5b-8 (13) and generation of the polymerized C9 responsible for MAC cytolytic activity (36,39,48). In contrast to CD59 that binds to C8␣ (30), paramyosin appears to bind both C8␣ and C8␤ chains (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…CD59 is an 18-kDa membrane glycoprotein expressed in most vertebrate tissues (9,10) that protects homologous cells from damage by complement. It binds to C8 and C9 (30,39) and inhibits assembly of the C5b-9n membrane attack complex (MAC) (36,48). As described here, sequencing and specific antibody binding data indicate that SCIP-1 is a tegumental form of paramyosin.…”

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Inhibition of the Complement Membrane Attack Complex bySchistosoma mansoniParamyosin

Deng¹,

Gold²,

LoVerde

et al. 2003

Infect Immun

100

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Following partial purification and analysis by mass spectrometry, we have determined SCIP-1 to be a surface-exposed form of the muscle protein paramyosin. As shown by immunofluorescence, anti-paramyosin antibodies label the surface of live schistosomula and adult worms. Like SCIP-1, purified native paramyosin reacts with a polyclonal rabbit anti-human CD59 antiserum, as shown by Western blot analysis. Also, the human complement components C8 and C9 bind to recombinant and native paramyosin. Analysis of paramyosin binding to fragments of C9 generated by thrombin or trypsin has demonstrated that paramyosin binds to C9 at a position located between Gly245 and Arg391. Paramyosin inhibited Zn 2؉ -induced C9 polymerization and poly-C9 deposition onto rabbit erythrocytes (E R ). In addition, paramyosin inhibited lysis of E R and of sensitized sheep erythrocytes by human complement. Finally, anti-paramyosin antibodies enhanced in vitro killing of schistosomula by normal and C4-depleted human complement. Taken together, these findings suggest that an exogenous form of S. mansoni paramyosin inhibits activation of the terminal pathway of complement and thus has an important immunomodulatory role in schistosomiasis.Schistosomiasis is one of the most prevalent parasitic diseases, affecting over 200 million people who live in areas of endemicity in Africa, South America, and Asia (12). Schistosoma mansoni, one of the causative agents of this disease, resides within patients' mesenteric and portal blood systems and successfully evades host immune responses (43). During skin penetration into the mammalian host and shortly afterwards, the larvae convert from exhibiting sensitivity to complement-mediated killing to bearing resistance to complementmediated killing (32, 49). The first step in that conversion is the removal of the glycocalyx coat that contains strong complement activators. Subsequently, the transformed schistosomula and the adult worms employ several strategies to evade the host's complement system (14), thus enabling the parasite to reside for years within the host's circulatory system in direct contact with complement proteins. This parasite contains proteins that bind in vitro to the complement proteins C1 (25) and C2 (22) and C8 and C9 (40) and may thus block the complement cascade at multiple steps. Paramyosin, one of the inhibitory proteins, was shown to bind in vitro to C1q and inhibit C1 and the classical pathway of complement activation (25). Paramyosin is in essence an invertebrate muscle protein (11) that serves as a major component of the thick filament and is thought to play an important role in certain specialized contractile states (7, 23). It has also been found to be an immunogen during infection of mice and humans with helminth parasites (27,37,47). A nonfilamentous membrane-bound form of paramyosin was also found in the tegumental outer layer (16, 33) and on the surface (18, 31) of S. mansoni schistosomes. Analyzed on the basis of its capacity to bind to Fc (31), it has been suggested to have immu...

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“…Based on the solved solution structure of CD59, these data suggest that selectivity for human C8 and C9 resides in a cluster of closely spaced side chains on the surface of CD59 contributed by His 44 CD59 is a 77-residue glycosylphosphatidylinositol (GPI) 1 anchored plasma membrane glycoprotein that serves to protect human blood and vascular cells from injury by the C5b-9 components of the complement system present in plasma (1)(2)(3)(4)(5)(6). The complement inhibitory function of CD59 resides in its capacity to interrupt formation of lytic pores in the plasma membrane by binding to segments of the C8␣ and C9 polypeptides that become exposed during their assembly into the complement C5b-9 membrane attack complex (MAC) (7)(8)(9)(10)(11)(12)(13). The amino acid sequence and solution structure of CD59 conform to those of the Ly6/cardiotoxin protein superfamily in which five internal disulfide bonds stabilize the polypeptide into a discoid structure consisting of a three-stranded ␤-sheet apposed to a two-stranded ␤-finger (14 -17).…”

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confidence: 99%

“…This implies that the CD59 homologue expressed on the surface of rabbit cells is also selective for homologous complement and cannot effectively interact with the human C8 and C9 components to inhibit MAC. We have previously taken advantage of the species selectivity inherent to the interaction of CD59 with C8 and C9 to map the peptide-binding sites within the human C8 and C9 proteins that are recognized by human CD59 (7,8,26). In those studies, the capacity of human CD59 to inhibit MAC assembly was measured using recombinant human/rabbit chimeras of C8 and C9 in order to identify the residues within the C8␣ and C9 polypeptides that are specifically required to confer recognition of these complement components by human CD59.…”

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confidence: 99%

Identity of the Residues Responsible for the Species-restricted Complement Inhibitory Function of Human CD59

Zhao

Zhou³

et al. 1998

Journal of Biological Chemistry

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The membrane-anchored glycoprotein CD59 inhibits assembly of the C5b-9 membrane attack complex (MAC) of human complement. This inhibitory function of CD59 is markedly selective for MAC assembled from human complement components C8 and C9, and CD59 shows little inhibitory function toward MAC assembled from rabbit and many other non-primate species. We have used this species selectivity of CD59 to identify the residues regulating its complement inhibitory function: cDNA of rabbit CD59 was cloned and used to express human/rabbit CD59 chimeras in murine SV-T2 cells. Plasma membrane expression of each CD59 chimera was quantified by use of a 5-TAG peptide epitope, and each construct was tested for its ability to inhibit assembly of functional MAC from human versus rabbit C8 and C9. These experiments revealed that the species selectivity of CD59 is entirely determined by sequence contained between residues 42 and 58 of the human CD59 polypeptide, whereas chimeric substitution outside this peptide segment has little effect on the MAC inhibitory function of CD59. Substitution of human CD59 residues 42-58 into rabbit CD59 resulted in a molecule that was functionally indistinguishable from native human CD59, whereas the complementary construct (corresponding residues of rabbit CD59 substituted into human CD59) was functionally indistinguishable from rabbit CD59. Based on the solved solution structure of CD59, these data suggest that selectivity for human C8 and C9 resides in a cluster of closely spaced side chains on the surface of CD59 contributed by CD59 is a 77-residue glycosylphosphatidylinositol (GPI) 1 -anchored plasma membrane glycoprotein that serves to protect human blood and vascular cells from injury by the C5b-9 components of the complement system present in plasma (1-6). The complement inhibitory function of CD59 resides in its capacity to interrupt formation of lytic pores in the plasma membrane by binding to segments of the C8␣ and C9 polypeptides that become exposed during their assembly into the complement C5b-9 membrane attack complex (MAC) (7-13). The amino acid sequence and solution structure of CD59 conform to those of the Ly6/cardiotoxin protein superfamily in which five internal disulfide bonds stabilize the polypeptide into a discoid structure consisting of a three-stranded ␤-sheet apposed to a two-stranded ␤-finger (14 -17). In CD59, the GPI anchor is attached to the C-terminal Asn 77 , and Asn 18 provides a single site of N-linked glycosylation (14,15,18). Whereas the carbohydrate attached to Asn 18 occludes most of one face of the discoid surface of the protein, these glycosidic residues do not contribute to the MAC inhibitory properties of CD59 (14, 15, 18 -20). This suggests that amino acid side chains exposed on the non-glycosylated face of the protein provide this function.The complement inhibitory function of CD59 exhibits marked selectivity for MAC assembled from C8 and C9 of human or primate origin, and CD59 shows little inhibitory function toward these same complement components of rabbi...

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Identity of the Segment of Human Complement C8 Recognized by Complement Regulatory Protein CD59

Cited by 57 publications

References 21 publications

Structure of Human C8 Protein Provides Mechanistic Insight into Membrane Pore Formation by Complement

Structure of Human C8 Protein Provides Mechanistic Insight into Membrane Pore Formation by Complement

Inhibition of the Complement Membrane Attack Complex bySchistosoma mansoniParamyosin

Identity of the Residues Responsible for the Species-restricted Complement Inhibitory Function of Human CD59

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