SUBJECT: IMMUNOLOGY & IMMUNOCHEMISTRY

MBIO 861

Complement System : Complement Proteins

Akinsemolu A, Department of Biosciences & Biotechnology, Babcock University, Ilishan-Remo, Nigeria Instructor Dr. Efuntoye Course: Immunology & Immunochemistry (MBIO 812)

Introduction
The complement system is a set of over 20 different protein molecules always found in the blood. However, a complement protein can be defined as a substance that is produced by a predecessor protein or in response to the presence of foreign material in the body and that triggers or participate in a complement reaction. Complement control proteins work in concert to regulate the system and keep it from damaging host tissue while simultaneously directing it towards foreign particles such as viruses and bacteria, and unwanted material such as cell debris and antibody-antigen complexes. (Janeway, et. al, 2001)

Members
The best-studied members of this family are: Complement receptor 1 (CR1 or CD35) Membrane cofactor protein (MCP or CD46) C4b-binding protein (C4BP). Decay-accelerating factor (DAF or CD55) Factor H (fH) (Source: Wikipedia.com)

Structure
RCA proteins typically formed by CCP domains, also termed Sushi domains or Short Consensus Repeats (SCR). Such beta-sandwich domains contain about 60 amino acid

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MBIO 861

residues each of that has 4 conserved cysteines, arranged in two conserved disulfide bonds (oxidized in 'abab' manner), and one also conserved tryptophan, but otherwise can vary greatly in sequence. The first CCP structure determined was a solution structure of the 16th module of factor H (pdb:1hcc).(Norman, 1991) Since then, other CCP domains have been solved either by NMR-spectroscopy (also relaxation studies, e.g. module 2 and 3 from CD55 (pdb:1nwv) (Uhrinova et.al, 2003) or by X-ray diffraction (also with co-crystallized partner, e.g. CR2 CCP modules complexed with C3d (pdb:1ghq)(Szakonyi, 2001).

Effects of the complement system/Proteins.

The main effect of the complement system is the induction of a pathogen-associated and modulated enzymatic cascade that, once triggered, ends with the lysis of the target cell and protects the host from infection. In addition to this apparent effect, the complement system also displays crucial additional activities that appear to be even more relevant.

One effect is the opsonization of the pathogen. Cleavage products such as C3b and C4b as well as C5b opsonize the surface of recognized pathogenic sub-stances and therefore facilitate phagocytosis. Additionally, opsonization is also important for the clearance of soluble, circulating antigen-antibody complexes. After the attachment of C3b and C4b to these complexes, they are bound to complement receptor 1 (CR1) on erythrocytes and are subsequently transported to the spleen and liver, where the immune complexes are eliminated.

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SUBJECT: IMMUNOLOGY & IMMUNOCHEMISTRY

MBIO 861

C3 cleavage products also bridge the innate and the adaptive immune systems. Opsonized antigens are bound to the complement receptor 2 (CR2) on B-cells via the C3-fragment C3d, initiating the production of specific antibodies as well as the differentiation of B-memory cells.

Presumably, the most important function is the induction of an anaphylactoid reaction. The small activation products C3a, C4a and particularly C5a are potent anaphylatoxins, capable of inducing the migration of phagocytes (Marder, 1985), smooth muscle relaxation, degranulation of mast cells and basophile granulocytes and therefore unleashing vasoactive substances such as histamine, prostaglandins, kinins and serotonin. All can cause vasodilation and capillary leakage (Schumacher, 1991) and induce the migrated cells to release eicosanoids, oxygen radicals and lysosomal enzymes, which cause damage to the pathogens (Goldstein, 1974; Mollnes, 2002; Sacks, 1978). It is noteworthy, that complement acts far beyond inflammation, as indicated by its close interaction with the coagulation cascade (Laudes, 2002; Amara, 2008; Markiewski, 2007) and its involvement in the regulation of apoptosis (Guo, 2000; Markiewski, 2009; Perianayagam, 2002; Riedemann, 2002) and cellular growth (Markiewski, 2008). The anaphylactoid functions are mediated by the interaction of C3a and C5a with their corresponding seven-transmembranespanning receptors C3aR and C5aR (CD88), respectively. The role of the second C5a receptor named C5L2 is not fully understood

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MBIO 861

and is still controversially discussed. However, there is increasing evidence that C5L2 represents a functional receptor acting as a negative regulator of the inflammatory response. For example, it was shown that inflammation in C5L2 knock-out mice was amplified (Gerard, 2005) and that blockage of C5L2 increased serum interleukin (IL)-6 (Gao, 2005).

In summary, complement is highly capable of inducing all classical signs of inflammation, with the occurrence of pain, swelling, reddening, hyperthermia and impaired function.
Complement Regulatory Proteins

Classical pathway Ag-Ab C1q C1INH C1rs MASP1, 2 MBL Microbes PNH C6, C7, C8, C9 CD59 S protein clusterin HAE MPGN

Alternative pathway

C4bp C4, C2 C4b2a DAF

C3

H C3bBb DAF, CR1

Neisseria meningitis D, P

C3b, B

Microbes

C3b + C5

H, MCP I

C3bi

CR3 LAD Bacterial infections

C5b-9 MAC

SC5b-9

Figure 1 An overview of complement activation, regulation and diseases caused by deficiencies of complement regulator proteins. The C1r and C1s are serine esterases that are of complement activation, regulation and diseases also inhibits analogous MBL-associated serine proteases proteins. Figure 2. An overviewinhibited by the plasma protein C1 inhibitor (C1 INH). C1 INHcaused by deficiencies of complement regulator(MASP-1 and MASP-2). Activity of the classical pathway C3/C5 convertase, C4b2a, is inhibited by the plasma factor C4b-binding protein (C4bp). The activity of the The C1r and C1s are serine esterases that are inhibited by the plasma protein C1 inhibitor (C1 INH). C1 INH also inhibits analogous alternative pathway C3/C5 convertase, C3bBb, can be enhanced by the only known physiological positive complement regulator, properdin (P). The selfMBL-associated serine proteases (MASP-1 and MASP-2). Activity of the classical pathway C3/C5 convertase, C4b2a, is inhibited by amplifying process of the AP is inhibited by multiple regulator molecules described in Figure 2. The five terminal plasma glycoproteins (C5, C6, C7, C8 and C9) bind sequentially to each other to generate the cytolytic The activity of the alternative pathway C3/C5 convertase, C3bBb, can terminal C the plasma factor C4b-binding protein (C4bp). membrane attack complex (MAC). Soluble regulators S protein and clusterin keep formingbe enhanced complexes in the fluid phase. On human cell membranes the main inhibitor of MAC is CD59 (protectin). Consequences of major complement regulator deficiencies by the only known physiological positive complement regulator, properdin (P). The self- amplifying process of the AP is inhibited are indicated by broken arrows. HAE, hereditary angio-oedema; PNH, paroxysmal nocturnal haemoglobinuria; MPGN, membranoproliferative glomerulonephritis; byLAD, leucocyte adhesion deficiency. multiple regulator molecules described in Figure 2. The five terminal plasma glycoproteins (C5, C6, C7, C8 and C9) bind sequentially to each other to generate the cytolytic membrane attack complex (MAC). Soluble regulators S protein and clusterin keep forming terminal C complexes in the fluid phase. On human cell membranes the main inhibitor of MAC is CD59 (protectin). Consequences of major complement regulator deficiencies are indicated by broken arrows. HAE, hereditary angio-oedema; PNH, paroxysmal nocturnal haemoglobinuria; MPGN, membranoproliferative glomerulonephritis; LAD, leucocyte adhesion deficiency.

shown to inhibit plasmin and coagulation factor XIa, but that C4bp irreversibly displaces C2a from C4b2a. The these functions are of less importance. Recent studies cofactor activity of C4bp leads to inactivation of C4b to indicate that C1 INH can also inhibit the MBL-associated C4c and C4d. serine proteases MASP-1 and MASP-2. PAGE 4 OF 6 COMPLEMENT SYSTEM: COMPLEMENTARY PROTEINS Factor H C4b-binding protein An analogue of C4bp in the alternative pathway is factor H C4b-binding protein (C4bp) is a multichain inhibitor of the (b1H-globulin). It is a uid-phase inhibitor of the AP C3

SUBJECT: IMMUNOLOGY & IMMUNOCHEMISTRY

MBIO 861

The functions of complement proteins can also be summarized: 1. Make bacteria more susceptible to phagocytosis 2. Directly lysing some bacteria and foreign cells 3. Producing chemotactic substances 4. Increasing vascular permeability 5. Causing smooth muscle contraction 6. Promoting mast cell degranulation (Douglas, 2010)

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COMPLEMENT SYSTEM: COMPLEMENTARY PROTEINS

SUBJECT: IMMUNOLOGY & IMMUNOCHEMISTRY

MBIO 861

References
Amara U, et al. (2008) Interaction between the coagulation and complement system. Adv. Exp. Med. Biol. 632:719. Douglas F.F. (2010) Components and Function of the Complement System Adv. Exp. Med. Biol. 632:719. Janeway C, Travers P, Wallport M, Schlomchik M. 2001. Immunobiology: The Immune System in Health and Disease. 5th ed. New York: Garland Publishing Gerard NP, etal. (2005) An anti-inflammatory function for the complement anaphylatoxin C5a-binding protein, C5L2. J. Biol. Chem. 280:3967780. Goldstein IM, Weissmann G. (1974) Generation of C5-derived lysosomal enzyme-releasing activity (C5a) by lysates of leukocyte lysosomes. J. Immunol. 113:158388. Guo RF, etal. (2000) Protective effects of anti-C5a Invest. 106:127180. Laudes IJ, etal. (2002) Anti-c5a ameliorates coagulation/fibrinolytic protein changes in a rat model of sepsis. Am. J. Pathol. 160:186775. Markiewski MM, Nilsson B, Ekdahl KN, Mollnes TE, Lambris JD. (2007) Complement and coagulation: strangers or partners in crime? Trends Immunol. 28:18492. Markiewski MM, etal. (2008) Modulation of the antitumor immune response by complement. Nat.m Immunol. 9:122535. Markiewski MM, etal. (2009) The regulation of liver cell survival by complement. J. Immunol. 182:541218. Mollnes TE, etal. (2002) Essential role of the C5a receptor in E coli-induced oxidative burst and phagocytosis revealed by a novel lepirudin-based human whole blood model of inflammation. Blood100:186977. Norman, D.G., Barlow, P.N., Baron, M., Day, A.J., Sim, R.B., Campbell, I.D. (1991) Three-dimensional structure of a complement control protein module in solution. J Mol Biol 219(4):717-725. Perianayagam MC, Balakrishnan VS, King AJ, Pereira BJ, Jaber BL. (2002) C5a delays apoptosis of human neutrophils by a phosphatidylinositol 3-kinase-signaling pathway. Kidney Int. 61:45663. Riedemann NC, etal. (2002) C5a receptor and thymocyte apoptosis in sepsis. FASEB J. 16:887-8. Sacks T, Moldow CF, Craddock PR, Bowers TK, Jacob HS. (1978) Oxygen radicals mediate endothelial cell damage by complement-stimulated granulocytes. An in vitro model of immune vascular damage. J. Clin. Invest. 61:116167. Schumacher WA, Fantone JC, Kunkel SE, Webb RC, Lucchesi BR. (1991) The anaphylatoxins C3a and C5a are vasodilators in the canine coronary vasculature in vitro and in vivo. Agents Actions 34:34549. Szakonyi, G., Guthridge, J.M., Li, D., Young, K., Holers, V.M., Chen, X.S. (2001) Structure of complement receptor 2 in complex with its C3d ligand. Science 292(5522):1725-1728.

Uhrnova, S., Lin, F., Ball, G., Bromek, K., Uhrin, D., Medof, M.E., Barlow, P.N. (2003) Solution structure of a functionally active fragment of decay-accelerating factor. Proc Natl Acad Sci USA 100(8):4718-4723.

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