TEMA 2. SISTEMA DE COMPLEMENTO - Historia - Componentes - Nomenclatura Proteins with assigned numbers, C1, C2... Many are zymogens, i.e. proenzymes which require proteolytic cleavage to become active. The enzymatically active form is distinguished from its precursor by a bar drawn above. The cleavage products of complement proteins are distinguished from parent molecules by suffix letters, C3a, C3b, etc. The proteins of the alternative pathway are called 'factors' and are identified by single letters, e.g. factor B, which may be abbreviated to FB or just 'B'.

Inmunologí Inmunología

Tema 2. El sistema del complemento

3. Funciones del sistema del complemento 1.

Intrinsic ability to lyse the cell membranes of many bacterial species.

2.

Complement products released in this reaction attract phagocytes to the site of the reaction - chemotaxis.

3.

Complement components coat the bacterial surface - opsonization allowing the phagocytes to recognize the bacteria and engulf them. Intrinsic ability of the complement system to recognize microbial components/antibodies bound to the microorganism.

4.

Binding of complement to receptors of immune cells - triggering inflammation, secretion immunorregulatory molecules…

5.

4.

5. Immune clearance- removal from immunocomplexes from circulation. Inmunologí Inmunología

Tema 2. El sistema del complemento

1

Both classical and alternative pathways generate a C3 convertase, which converts C3 to C3b, the central event of the complement pathway. C3b in turn activates the terminal, lytic sequence, C5-C9. The first stage in the classical pathway is the binding of antigen to antibody. The alternative pathway does not require antibody and is initiated by the covalent binding of C3b to hydroxyl and amine groups on the surface of various microorganisms. The alternative pathway provides nonspecific 'innate' immunity, whereas the classical pathway represents a more recently evolved link to the adaptive immune system.

Inmunologí Inmunología

Tema 2. El sistema del complemento

Vía clásica The classical pathway is activated by the cleavage of C1r and C1s following association of C1qr2s2 with classical pathway activators including immune complexes. Once activated cleaves C4 and C2 to form the classical pathway C3 convertase.

Inmunologí Inmunología

Tema 2. El sistema del complemento

2

C1, primer componente de la vía clásica Electronmicrograph of a human C1q molecule demonstrates six subunits. Each subunit contains three polypeptide chains, giving 18 in the whole molecule. The receptors for the Fc regions of IgG and IgM are in the globular heads. The connecting stalks contain regions of triple helix and the central core region contains collagen-like triple helix. Model of intact C1 with two C1r and two C1s proenzymes positioned within the ring. The catalytic heads of C1r and C1s are closely apposed and conformational change induced in C1q following binding to complexed immunoglobulin causes mutual activation/cleavage of each C1r unit followed by cleavage of the two C1s units. The cohesion of the entire complex is dependent on Ca2+.

Inmunologí Inmunología

Cabeza globular

Conector triple hélice núcleo collagen-like

Tema 2. El sistema del complemento

Unión Fc de IgG a dominio globular de C1q Cambio conformacional C1 C1r (1) autocatálisis C1r (2) activación C1s (1 y 2) activación- serinesterasas activas sobre C4 y C2 Formación C4b2a- C3 convertasa de la vía clásica

3

Vía de las lectinas Cleavage of C4 and C2 can also be effected via the lectin pathway, which is associated with mannanbinding lectin (MBL).

Inmunologí Inmunología

Tema 2. El sistema del complemento

Activadores de las vías clásica y de las lectinas

Inmunologí Inmunología

Tema 2. El sistema del complemento

4

Vía alternativa The alternative pathway is activated by the cleavage of C3 to C3b, which associates with factor B and is cleaved by factor D to generate the alternative pathway C3 convertase C3bBb. The initial activation of C3 happens to some extent spontaneously. This step can also be effected by classical or alternative pathway C3 convertases or a number of other serum or microbial proteases. Activation pathways are functionally analogous. C3 and C4 are homologous, as are C2 and factor B. MASP-1 and MASP-2 are homologous to C1r and C1s, respectively.

Inmunologí Inmunología

Tema 2. El sistema del complemento

Activación del enlace tioester de C3

The α-chain of C3 contains a thioester bond formed between a cysteine and a glutamine residue, with the elimination of ammonia. Following cleavage of C3 into C3a and C3b*, the bond becomes unstable and susceptible to nucleophilic attack by electrons on -OH and -NH2 groups, allowing the C3b to form covalent bonds with proteins and carbohydrates - the active group decays rapidly by hydrolysis, if such a bond does not form. C4 also contains a thioester bond which becomes activated similarly when C4 is split into C4a and C4b. Inmunologí Inmunología

Tema 2. El sistema del complemento

5

Activación / asa de amplificación / regulación de la vía alternativa

Alternative pathway activation depends on the presence of protected surfaces. Amplification: 'Protected' means that bound C3b is protected from proteolytic degradation. C3b bound to an activator surface binds factor B, which is cleaved by factor D to produce the alternative pathway C3 convertase , which drives the amplification loop, by cleaving more C3- higher C3 deposition Inhibition: On self surfaces the binding of factor H is favoured and C3b is inactivated/catabolised by factor I. The binding of factor B or factor H controls the development of the alternative pathway reactions. CR1 and DAF limit complement activation on self cell membranes - cofactors of factor I in dissociation of C3bBb convertase. Protected surfaces: bacterial immunoglobuline aggregates.

Inmunologí Inmunología

membranes,

Tema 2. El sistema del complemento

Inactivación de C3

Factor I cleaves C3b in three places to release C3c, leaving C3dg, a fragment of the α-chain, still bound to the substrate. The first two cleavages are promoted by factor H, MCP or CR1 and produce an intermediate iC3b. The third cleavage is promoted by CR1.

Inmunologí Inmunología

Tema 2. El sistema del complemento

6

Activadores de la vía alternativa

Inmunologí Inmunología

Tema 2. El sistema del complemento

Vía lítica

Classical (C4b2a) or alternative (C3bBb) pathway C3 convertases associate with C3b bound on a cell surface to form C5 convertases (C4b2a3b / C3bBb3b), which split C5. The larger fragment, C5b associates with C6 and C7, which can then bind to plasma membranes. The complex of C5b67 assembles C8 and a number of molecules of C9 to form a membrane attack complex (MAC), C5b-9.

Inmunologí Inmunología

Tema 2. El sistema del complemento

7

Vía lítica

Assembly of the C5b-9 membrane attack complex (MAC). Recruitment of a further C3b into the C3bBb enzymic complex generates a C5 convertase which cleaves C5a from C5 and leaves the remaining C5b attached to the membrane. Once C5b is membrane bound, C6 and C7 attach themselves to form the stable complex, C5b67, which interacts with C8 to yield C5b678. This unit has some effect in disrupting the membrane, but primarily causes the polymerization of C9 to form tubules traversing the membrane. The resulting tubule is referred to as a MAC. Disruption of the membrane by this structure permits the free exchange of solutes, which is primarily responsible for cell lysis Inmunologí Inmunología

Tema 2. El sistema del complemento

Reguladores de la activación del complemento

Inmunologí Inmunología

Tema 2. El sistema del complemento

8

Inmunologí Inmunología

Tema 2. El sistema del complemento

Las funciones del sistema del complemento

The complement system has an intrinsic ability to lyse the cell membranes of many bacterial species. (2) Complement products released in this reaction attract phagocytes to the site of the reaction - chemotaxis. (3) Complement components coat the bacterial surface - opsonization allowing the phagocytes to recognize the bacteria and engulf them. These reactions may be triggered by the intrinsic ability of the complement system to recognize microbial components or by antibodies bound to the microorganism.

Inmunologí Inmunología

Tema 2. El sistema del complemento

9

Opsonización

Upper panel: A bacterium is sensitized by the covalent binding of C3b, iC3b and C4b, which allow it to be recognized by complement receptors (CR) on neutrophils and mononuclear phagocytes. This promotes phagocytosis and activation of the phagocyte. In primates, erythrocytes also express CR1, which allows them to bind opsonized bacteria and immune complexes. In the lower panel fluoresceinated bacteria which have been opsonized with antibody and complement are seen adhering to human erythrocytes.

Inmunologí Inmunología

Tema 2. El sistema del complemento

Receptores del complemento

The complement receptors CR1 and CR2 are formed from numerous repeated complement control protein (CCP) domains. CR3 and CR4 are integrins which share a common β chain (CD18). CR1, CR3 and CR4 are expressed on macrophages and other phagocytic cells, while CR2 is confined to B cells and follicular dendritic cells, where it has a function in the maturation of the antibody response Inmunologí Inmunología

Tema 2. El sistema del complemento

10

Quimiotaxis

C5a and C3a both act on mast cells to cause degranulation and release of vasoactive amines, including histamine and 5-hydroxytryptamine, which enhance vascular permeability and local blood flow. The secondary release of chemokines from mast cells causes cellular accumulation and C5a itself acts directly on receptors on monocytes and neutrophils to induce their migration to sites of acute inflammation and subsequent activation.

Inmunologí Inmunología

Tema 2. El sistema del complemento

The table lists major inflammatory mediators, which control blood supply and vascular permeability or modulate cell movement. The main sources are given (centre block).

Inmunologí Inmunología

Tema 2. El sistema del complemento

11