The Complement System

Innate vs Adaptive Immunity

Antibodies

Antibodies: Soluble polypeptide molecules manufactured and secreted by B cells and plasma cells that can recognize any substance. They have neutralizing or cytotoxic activities (cell or complemented-mediated)

• Antibodies participate in both innate and adaptive immunity

  

• Structure of Antibodies and B cell receptors:

  

Antibody molecules can be cleaved into functionally distinct fragments

• Pepsin Digestion: The enzyme pepsin cleaves below the hinge region of the antibody. It generates a F(ab’)2 fragment (Fab arms still connected via disulfide bonds) and destroys Fc

• Papain Digestion: cleaves above the hinge region. It generates two separate Fab fragments and one intact Fc fragment.

• Mercaptoethanol Reduction: This reducing agent breaks disulfide bonds. It separates all the polypeptide chains

Six Effector Mechanisms of Antibodies

1. Neutralization of pathogens and toxins: Antibodies bind to pathogens inactivating them, preventing them from binding to cells.

2. Agglutination of particulate antigens: Prevents binding to cells, enhances clearance. Antibodies clump pathogens together making them easier to clear

3. Opsonization: Antibodies coat the surface of pathogens marking them for phagocytosis.

4. Complement activation: Antibodies trigger the classical complement pathway, leading to pathogen lysis, inflammation. etc.

5. Antibody-Dependent-Cell mediated Cytotoxicity (ADCC): Antibodies bound to infected cells interact with Fc receptors on NK cells which induce apoptosis

6. Degranulation: Antibodies bind to pathogens and trigger the release pre-stored granules which contain histamine, cytokines and enzymes

The complement system

The complement system: Components of plasma that complement antibody action to kill target cells by lysis. They are around 50 proteins that interact in a network as well as with other systems.

Complement functions include:

1. Alternative pathway: Directly kill cells

2. Classical pathway: Interact with antibodies to kill cells

3. Lectin pathway: Interact with lectins to kill cells

4. Opsonize pathogens to flag them for phagocytosis

5. Function as a chemoattractant to recruit leukocytes and induce inflammation

Immunoglobins and whether they activate the complement pathway:

Nomenclature:

• C: proteins that belong in the Classical pathway

• #: name of the component in order of discovery

• b: designates the larger fragment with enzyme activity

• a: designates the smaller fragment

Classical Pathway

• Trigger: Initiated by the pathogen sensor C1 complex binding to pathogen bound antibodies like IgG or IgM

• Goal: To form the membrane attack complex (MAC) and kill pathogens through lysis or opsonization

Steps of the Classical Pathway:

1. Initiation by C1 complex:

   • C1 Complex components:

     • C1q: 18 polypeptide chains in 6 collage like triple helices

     • C1r and C1s: 2 molecules each

     • Tails: Interact with 2 C1r and 2 C1s

     • Heads: Interact with IgM or IgG

       

   • C1q Activation:

     • C1q binds to antigen-bound antibody, ( IgM (staple form) or IgG on pathogen surfaces).

       

     • This induces a conformational change activating C1r, which then activates the second C1r and the two C1s molecules ….(C1r → 2nd C1r and 2 C1s)

2. Activation of C4 and C2:

   • C1s Activity:

     • Cleaves C4 into C4a (small) and C4b (larger)

     • C4b binds to pathogen surface

       

   • C2 binding and Cleavage:

     • C2 binds to C4b and is broken by C1s into C2a (larger) and C2b (smaller and not on the bacteria anymore)

       

     • C4b2a complex forms and acts as the C3 convertase

       

3. Formation of C3 Convertase:

   • C4b2a Complex

     • breaks C3 into C3a (small fragment, chemoattractant, not on the complex) and C3b (larger fragment, in the complex)

       

     • C3b coats the pathogen surface for opsonization and amplifies the complement cascade

4. Formation of C5 Convertase:

   • C4b2a3b Complex:

     • Formed when one C3b molecule binds to C4b2a

     • Functions as the C5 Convertase

   • C5 cleavage:

     • Breaks C5 into C5a and C5b ?????

Diagram/Flow (for Quick Review)

1. C1 Activation: C1q binds IgM/IgG → Activates C1r → Activates C1s.

2. C4 Cleavage: C1s breaks C4 → C4b binds pathogen.

3. C2 Cleavage: C4b binds C2 → C2 cleaved to form C4b2a (C3 convertase).

4. C3 Cleavage: C3 convertase cleaves C3 → C3b coats pathogen; amplifies cascade.

5. C5 Convertase: C4b2a3b cleaves C5 → Leads to MAC formation.

Lectin Pathway

Trigger: MBL or ficolins bind to polysaccharide antigen on the pathogens surface

Goal:

Steps:??

• MBL/ficolin binds pathogen surface.

• Activated MASPs cleave C4 and C2.

• Formation of C3 convertase (C4b2a) → Cleaves C3 into C3a and C3b.

• C3b opsonizes pathogens and continues the complement cascade.

  

Alternative pathway

Trigger: Spontaneous cleavage of C3 into C3a and C3b. Requires no antibodies, lectins or pathogen-bound triggers.

Goal: form MAC complex

Steps of Alternative Pathway:

1. Spontaneous Cleavage

   • C3 undergoes spontaneous hydrolysis to from C3b.?

2. Formation of C3 Convertase

   • C3b binds Factor B, which is cleaved by Factor D to form C3bBb (a fluid phase C3 convertase)

     

   • C3 is broken down into C3b and C3a(leaves) and C3b attaches the pathogen

     

3. Stabilization:

   • Properdin stabilizes the C3bBb complex on microbial surfaces

4. C5 Convertase formation:

   • Addition to another B forms C3b, making C3bBbC3b, which acts as a C5 convertase. Required Factor D y Properdin

     

5. C5 cleavage:

   • C5 convertase cleaves C5 breaks into C5a and C5b → Leads to MAC formation.

     

     *C3a and C5a are potent inflammatory signals

Formation of the Membrane attack complex (MAC)

Regulation of Complement

• Mechanism to prevent complement attack on self-cells:

  1. Unstable complement enzymes

  2. C1 inhibitor (C1NH): Dissociates C1q from C1s/C1r

  3. Decay-accelerating factor (DAF) and complement receptor 1 (CR1): Dissociate C3 convertases

Genetic deficiencies in complement

• C3 Deficiency: Susceptibility to bacterial infections

• C5-C9 Deficiency: Recurrent infections with Neisseria species

• C1q or C4 Deficiency: Linked to systemic lupus erythematosus (SLE)