Complement System

Discovery
• Jules Bordet (Pasteur Institute, Paris)
– Sheep antiserum against V. cholerae cause lysis of bacteria.
– When serum was heated, the ability to cause lysis was lost.
– Addition of fresh serum (lacking Abs directed against
bacterium) to the heated serum, ability of lysis generated
again.
• Bordet reasoned – Antibacterial activity requires two
components
– Abs
– Heat sensitive component
• Paul Ehrlich (Berlin) – Similar experiments – Coining of
term Complement.
 Complement
• Activation by Abs
• Components of Innate Immunity
 Components of Complement
• Complement proteins (soluble proteins and
glycoproteins)-
– Synthesized mainly by liver hepatocytes.
– Significant amounts also produced by
• blood monocytes
• tissue macrophages
• epithelial cells of gastrointestinal and genitourinary
tract
• Circulate in the functionally inactive forms
(proenzymes, zymogens)
 Components of Complement
• Complement components designated by
– numerals C1 - C9
– Letter symbols – factor D
– Trivial names – homologous restriction factor
• Peptide fragments formed by activation of
component is denoted by small letters (C3 –
C3a, C3b). Large fragment denoted by ‘b’ and
small fragment by ‘a’. Opposite in case of C2 –
C2a – large fragment, C2b – small fragment.
 Components of Complement
• Complement reaction sequence starts with an
enzyme cascade.
• Cleavage of complement protein generates a
larger fragment and smaller fragment
– Large fragment – binds to the target near the site
of activation
– Small fragment – diffuse from the site and initiate
localized inflammatory responses by binding to
specific receptors
 Components of Complement System
• Complement fragments interact to form
complexes. Those complexes that have
enzymatic activity are designated by a bar
over the number or symbol
– C4b2a
– C3bBb
 Complement Activation Pathways

Classical Alternative Lectin

C5b

Membrane Attack Complex (MAC)

 C1 Component
• C1 is composed of
– C1q – Composed of 18 polypeptide chains that
associate to form 6 collagen like triple helical
arms, the tips of which binds to CH2 domain of Ab
molecule
– C1r and C1s – have a catalytic domain and an
interaction domain.
 Classical Pathway
• Ab (IgM and IgG1, IgG2, IgG3)
– Formation of Ag-Ab complex
– Ab binding to a target site
• Formation of Ag-Ab complex induce
conformational changes in Fc portion of the
IgM molecule that expose binding site for C1
component. The C1 component consists of 1
C1q and 2 molecules of each C1r and C1s
forming a complex C1qr2s2 - stabilized by Ca2+
 Classical Pathway
• Each C1 macromolecular complex must bind
by its C1q globular heads to at least two Fc
sites of Ab. Pentameric IgM when binds to the
Ag it assumes a staple configuration in which
C1 binding sites are exposed but when in
circulation it assumes planar configuration in
which C1 binding sites are not exposed. Due
to its pentameric form IgM is more efficient in
complement activation.
Planar Form
Staple Form
 Classical Pathway
• Binding of C1q to Ab induce conformational
changes in C1r converting it to C1r (serine
protease) which then cleaves C1s to similar
enzyme C1s
• C1s of C1qr2s2 acts on C4 and C2.
• C4 is a glycoprotein composed of three chains
(,,). - chain is cleaved exposing a binding
site on C4b
 Classical Pathway
• C4b attaches to the target surface in vicinity of C1.
• C2 proenzyme then binds to C4b, where C2 is
cleaved by C1s forming C4b2a (C3 convertase). C2b
diffuses away
• C4b2a (C3 convertase) act on C3 (made up of  & 
chains) and cleaves a part of  chain generating
C3b and C3a. This is an amplification step in
complement activation. A single C3 convertase
molecule can generate 200 molecules of C3b.
 Classical Pathway
• Some C3b binds to C4b2a forming C4b2a3b
(C5 convertase)
• C5 binds to C3b of C4b2a3b complex. This
binding alters conformation of C5 and it is
then cleaved by C4b2a portion of C5
convertase to produce C5b and C5a.
 Classical Pathway
Ag+Ab

Activates C1. Binding of C1q to Fc portion of Ab.

Induce conformational changes in C1r converting it to

C1r (serine protease) which then cleaves C1s to
similar enzyme C1s

C1s acts on C4. - chain of C4 is cleaved to C4b,

which binds to the target in vicinity to C1 complex
C4a
 C2 binds the exposed C4b, where it is cleaved by
C1s forming C4b2a (C3 convertase)
C2b
C3( chain) is cleaved by C3 convertase forming
C3b and C3a. This step is amplification step
C3b, C3a
C3b joins C4b2a forming C4b2a3b (C5 Convertase)

C4b2a3b (C5 Convertase) acts on C5 forming C5b

C5a
C5b
 Alternative Pathway
• Antibody Independent
• C3 component containing unstable thioester bond
undergo spontaneous hydrolysis to yield C3a and C3b.
• C3b binds to the foreign surface Ags
• C3b present on foreign Ag binds to Factor B. The
complex of C3b and Factor B is stabilised by Mg2+.
• Factor D cleaves C3b bound factor B releasing Ba (small
fragment) which diffuses away and generating C3bBb
(C3 convertase)
• C3bBb is unstable having limited half life and is
stabilized by the binding of properdin.
 Alternative Pathway
• C3bBb further cleaves C3 producing C3b
(amplification) which joins the complex
C3bBb3b (C5 convertase).
• Nonenzymatic component C3b binds C5 and
Bb component hydrolyzes it producing C5b
 Lectin Pathway (MB Lectin/Mannan
Binding Lectin)
• Antibody Independent
• Initiated by binding of Mannose Binding Lectin (MBL)
to the mannose residues on the surface of
microorganism (bacteria, fungi, viruses)
• MBL is an acute phase protein and a member of
collectin family. MBL resembles C1q in structure and
function.
• Following this MBL Associated Serine Protease (MASP)
1 and 2 bind to MBL. MASP-1 and MASP-2 are similar
to C1r and C1s structurally and mimics their activity.
• MASP-1 and MASP-2 activate C4 and C2 forming C4b2a
 Comparison
Property Classical Alternative Lectin
Antibody Yes No No
Mediated

Initiated by C1 binding to Spontaneous Mannose

Ab cleavage of Binding
C3 Lectin
C3 C4b2a C3bBb C4b2a
Convertase
C5 C4b2a3b C3bBb3b C4b2a3b
Convertase
Similarity in C1q - MBL
C1r - MASP-1
C1s - MASP-2
 Membrane Attack Complex (MAC)
• C5b binds to the surface of target cell, (but it is
extremely labile and degrades rapidly)
• It is stabilized by the binding of C6 forming C5b6
• C5b6 binds to C7 and undergoes structural transition
exposing the hydrophobic regions. C5b67 inserts in the
membrane of the target cell. In case of Ag-Ab complex
it is released and may bind to other cells in vicinity
leading to ‘innocent bystander’ lysis
• Binding of C8 to C5b67 takes place resulting in
conformational change in C8 exposing the hydrophobic
region which interacts with plasma membrane. C5b678
o
causes a 10A pore in the membrane.
 MAC
• Finally binding and polymerization of C9
(perforin like molecule) takes place. During
polymerization C9 undergoes transition so
that it can be inserted in the membrane. 10-
17 molecules of C9 can be bound to a single
C5b678 complex.
• MAC consists of poly C9 surrounding the
C5b678 complex. This creates pores of 70-
100A .
o
MAC
 Biological Consequences of
Complement Activation
• Lysis of cells, bacteria, viruses
• Opsonization leading to phagocytosis
• Binding of specific complement receptors on
cells of immune system triggering specific cell
functions
• Clearance of immune complexes from
circulation
 Biological Consequences of
Complement Activation
• MAC - lysis of cells
• C3a, C4a, C5a – Anaphylotoxins –
Degranulation of mast cells and basophils
• C3a, C5a – degranulation of eosinophils
• C3a, C5a, C5b67 – Extravasation and
chemotaxis of leukocytes
• C3a, C5a – Aggregation of Platelets
 Biological Consequences of
Complement Activation
• C5a – Release of hydrolytic enzymes from
neutrophils and increase in expression of
complement receptors type 1 and 3 (CR1, CR3)
on neutrophils
• C3b, C4b, – Opsonization and phagocytosis
• C3b, MAC – Virus neutralization
• C3b – Clearance of Immune Complexes