Complement System Overview

The Complement System

• A defensive system consisting of over 30 proteins (activators and regulators) produced by the liver and found in circulating blood serum
• Discovered by Jules Bordet in 1896
• Inactivated in the lab by heating serum at 56 degrees Celsius for 30 minutes

Complement System Nomenclature

• Beta-1 and Beta 2 globulins are fractions of serum proteins separated by electrophoresis
• Complement proteins are included in these globulins, especially C3 and C4
• Complement proteins are named with a capital "C", followed by a number (e.g., C3)
• A small letter after the number indicates the size of the protein after activation
• 'a' indicates a smaller fragment that diffuses away from the surface
• 'b' indicates a larger fragment that binds to the cell surface
• Nomenclature is based on order of discovery, NOT order of activation (C1, C4, C2, C3)

Classical Pathway

• Considered part of the specific/adaptive immune response as it relies on antibodies
• Activators:
  • Antigen-antibody complex
  • Apoptotic cells
  • Certain viruses / gram-negative bacteria
  • CRP bound to ligand
• Initiated when C1 binds to the ends of antibodies (at least 2 CH2 domains)
• IgM is more effective at complement fixation than IgG

Recognition Unit (Classical Pathway)

• C1 complex is the recognition unit of the Classical Pathway
• C1 recognizes at least 2 CH2 domains of antibodies
• C1 has 3 subunits: C1q, C1r, C1s
• C1q recognizes 2 CH2, activates C1r, which in turn activates C1s

C3 Activation Complex (Classical Pathway)

• Activated C1 activates C2 and C4
• C2 is cleaved into C2a and C2b
• C4 is cleaved into C4a and C4b
• C2b and C4b bind together on the bacterial surface, forming the C3 convertase
• C2a and C4a diffuse away
• The C3 convertase cleaves C3 into C3a and C3b
• This is the pivotal point where all 3 pathways converge
• All pathways lead to C3 cleavage, forming C5 convertase

C3b

• Many C3b molecules are produced
• C3b binds and coats the bacterial surface, promoting opsonization
• C3b is an opsonin – molecules that bind to both bacteria and phagocytes, increasing phagocytosis by 1,000 fold or more

C3a, C4a, C5a

• C3a, C4a, C5a are small fragments that diffuse away from bacteria
• These act as anaphylatoxins, triggering inflammation
• C3a binds to mast cells, causing histamine release
• C5a is the most powerful chemotactic factor known for leukocytes

C5 Activation Complex (Classical Pathway)

• The C5 convertase (C4b2b3b) cleaves C5 into C5a and C5b, which initiates complement-mediated cell lysis
• C5b binds to C6, initiating the formation of the Membrane Attack Complex (MAC)

Membrane Attack Complex (MAC)

• C5b binds to C6, activating C6 which binds to C7
• C7 binds to C8, which binds to C9s
• These proteins assemble to form a circular complex, the Membrane Attack Complex (MAC)
• MAC causes cytolysis by creating a transmembrane channel
• Cell contents leak out, cell integrity is compromised, cell death occurs
• Na+ ions and water also rush into the cell.

The Alternative Pathway

• Part of the non-specific/innate defense
• Activated by fungal cell wall (zymosan), snake venom, lipopolysaccharide, bacterial polysaccharides, and tumor cells
• Activated spontaneously on microbial surfaces without antibodies

Factor B (Alternative Pathway)

• C3b on the bacterial surface binds to Factor B
• C3b + Factor B = C3bBb
• Factor B resembles C2 in the classical pathway

Factor D (Alternative Pathway)

• Factor D cleaves Factor B into Ba and Bb
• Bb remains bound to C3b, while Ba and Factor D disperse
• The C3bBb complex is stabilized by Properdin (Factor P), forming the C3 convertase
• C3bBbP makes up the critical C3 activation complex in the alternative pathway

C3 Activation Complex (Alternative Pathway)

• The C3 convertase (C3bBbP) produces more C3b, amplifying the pathway
• This allows repeated steps in the pathway

C5 Activation Complex (Alternative Pathway)

• An additional C3b binds to the C3 convertase, converting it into the C5 convertase
• The C5 convertase cleaves C5 into C5a and C5b, initiating MAC formation

Classic vs. Alternative Pathways

• Classical Pathway:
  • Specific, acquired immunity
  • Initiated by antibody
  • Involves all components (C1, C4, C2)
  • Properdin system is NOT involved
• Alternative Pathway:
  • Non-specific, innate immunity
  • Initiated by microbial surfaces
  • C1, C4, and C2 are bypassed
  • Properdin system IS involved

Mannose Binding Lectin (MBL) Pathway

• Part of the non-specific, innate immunity
• MBL is a protein present in humans that binds to mannose on microbial surfaces
• Mannose is a sugar absent in humans but present in microbes
• MBL is structurally similar to C1q.
• MBL activates MBL-associated serine proteases (MASPs; MASP1 and MASP2), initiating the complement pathway
• MASP activation leads to C2 and C4 activation

Biological Functions of the Complement System

• Lysis of target cells: MAC
• Opsonization: C3b
• Anaphylatoxins: C3a, C4a, C5a (trigger inflammation)
• Increased vascular permeability: C2a
• Chemotaxis: C5a (recruits neutrophils)

Complement Regulators

• C1 Inhibitor (C1 INH): Prevents spontaneous activation of the classical pathway.
• C4b-Binding Protein (C4b-BP): Prevents C4b and C2b binding.
• Decay Accelerating Factor (DAF): Dissociates C3 convertases in the classical, lectin, and alternative pathways.
• Factor I: Cleaves C4b into smaller fragments (C4c and C4d), and also inactivates C3b and C4b; removing them from the cascade
• CD59: Prevents association of C5b678 with C9.
• Factor H: Prevents binding of C3b to Factor B, cleaves C3b into C3c and C3d.

Diseases and Disorders Associated with Complement Component Deficiencies

• C1 Inhibitor Deficiency: Hereditary Angioedema
• DAF Deficiency: Paroxysmal Nocturnal Hemoglobinuria
• MAC Deficiency: Recurrent infections (especially Neisseria meningitidis)

Total Complement Activity (CH50, CH100)

• Measures the integrity of the classical complement pathway (screening test)
• Based on the ability of serum to lyse sheep RBCs coated with anti-sheep antibodies
• One CH50 unit is defined as the volume or dilution of serum that lyses 50% of erythrocytes in the reaction mixture.