THE COMPLEMENT SYSTEM

(A COMPLEX INNATE IMMUNE SURVEILLANCE SYSTEM)

Subject : Zoology Prepared by : Neha Sharma

Paper : CBCZ -203: Immunology SOS in Zoology
Unit : III
CONTENTS :
 History
 Introduction
 Complement plays a key role in both Innate and Adaptive Immunity
 The Complement Components
 The Functions of Complement
 Three Pathways for Complement activation
 Complement Activation
 A Cascade System
 Cascade Activation
 The Classical Pathway
 The Alternative Pathway
 The Lectin Pathway (MAL Pathway)
 The 3 Complement Pathways converse at the “Membrane-Attack Complex
(MAC)”
 Regulation of Complement System
 Clinical aspects of Complement.
 HISTORY
Research on complement began in the 1890s, when
Jules Bordet (Institute Pasteur in Paris) observed:
1. Sheep antiserum to the bacterium Vibrio cholerae
caused lysis of the bacteria.
2. Heating the antiserum destroyed its bacteriolytic
activity.
3. Addition of fresh normal serum, that contained
no Abs against the bacterium and was unable to
kill the becterium by itself, restored the ability to
lyse the bacteria by the heated antiserum.
 Bacteriolytic activity requires 2 different substances.
a. The specific antibacterial Abs, which are resistant to the
heating process.
b. Heat-sensitive component responsible for the lytic activity.
 Paul Ehrlich in Berlin carried out similar experiments
and named the substance complement, defining it
as “the activity of serum that completes the action
of Ab.”
 Bordet won the Nobel Prize in 1919 –
“Complement-mediated bacteriolysis”
COMPLEMENT PLAYS A KEY ROLE IN BOTH
INNATE AND ADAPTIVE IMMUNITY

 The complement system is the major effector

of the humoral immune system.
 Although the discovery of complement and
most early studies were linked to the activity of
complement following Ab binding, a major role
for this system is the recognition and
destruction of pathogens based on recognition
of pathogen-associated molecular patterns, or
PAMPs, rather than on Ab specificity.
THE COMPLEMENT COMPONENTS
 More than 30 soluble and cell-bound protein.
 Participate in both innate and adaptive immunity.
 Produced by hepatocytes (mainly), monocytes and
epithelial cells of the gastrointestinal and
genitourinary tracts.
 Constitute 5% (by weight) of the serum globulin
fraction.
 Many components are proenzymens (zymogens),
which are functionally inactive until proteolytic
cleavage, which removes an inhibitory fragment and
exposes the active site.
 Reaction starts with an enzyme cascade.
THE FUNCTIONS OF COMPLEMENT
1. Cytolysis :
 Lysis of cells, bacteria, and viruses – the major
effector of the humoral branch of the immune
system.
 Disrupt the membrane & the entry of water and
electrolytes into the cell.
2. Opsonization :
• C3b & C1q; promtes phagocytosis of particulate Ags.
• Extremely important when pathogen carries a capsule.

C3b is an opsonin
Opsonin are
molecules that bind both
to bacteria and
phagocytes
Opsonin increses
phagocytosis by 1,000
fold.
3. Activation of Inflammatory Response :
Binding to specific complement receptors
on cells of the immune system, triggering
specific cell functions, inflammations, and
secretion of immunoregulatory molecules.
4. Clearance of Immune Complexes :
Immune clearance, which removes immune
complexes from the circulation and deposits
them in the spleen and liver.
THREE PATHWAYS FOR COMPLEMENT
ACTIVATION :
 1. Classical Pathway
 2. Alternative Pathway
 3. Lectin or MBL (mannose-binding lectin) Pathway.

 Pathway for activation require multiple steps

categorized as :
 1. Recognition

 2. Enzyme activation

 3. Biological activity
COMPLEMENT ACTIVATION

1. Classical Pathway – Begins with the formation of Ag-

Ab complex.

2. Alternative Pathway – is initiated by cell-surface

constituesnts that are foreign to the host.
- Ab-independent.

3. Lectin Pathway – is activated by the binding of

mannose-binding lectin (MBL) to mannose residues
on glycoproteins or carbohydrates on the surface of
microorganisms.
- Ab-independent.
A CASCADE SYSTEM

 The complement works as a cascade system.

- Cascade is when one reaction triggers another
reaction which triggers others and so on. These types
of systems can grow exponentially very fast.
CASCADE ACTIVATION

 Complements proteins are often designated by

an uppercase letter C and are inactive until
they are split into products.
- Example: C1
 When the products are split they become
active. The active products are usually
designated with a lowe case a or b.
- Example: C1a and C1b
THE CLASSICAL PATHWAY

 The classical pathway

is considered to be
part of the specific
immune response
because it relies on
antibodies to initiate it.
 C1 becomes activated
when it binds to the
ends of antibodies.
THE BUILDING OF A C3 ACTIVATION COMPLEX

 Once C1 is activated, it
activates 2 other
complement proteins, C2
and C4 by cutting them in
half.
 C2 is cleaved into C2a and
C2b.
 C4 is cleaved into C4a and
C4b.
 Both C2b and C4b bind
together on the surface of
the bacteria.
 C2a and C4a diffuse away.
 C3 ACTIVATION COMPLEX

 C2b and C4b bind

together on the surface
to form a C3 activation
complex.
 The function of the C3
activation complex is to
activate C3 proteins.
- This is done by cleaving C3
into C3a and C3b.
C3A

C3a increase the inflammatory response by binding to

mast cells and causing them to release histamine.
 C3B

 Many C3b molecules are produced by the C3

activation complex.
 The C3b bind to and coat the surface of the
bacteria.
 C3b is an opsonin :
- Opsonins are molecules that bind both to bacteria and
phagocytes.
- Opsonization increases phagocytosis by 1,000 fold.
BUILDING THE C5 ACTIVATION COMPLEX
 Eventually enough C3b is cleaved that the
surface of the bacteria begins to become
saturated with it.
 C2b and C4b which make up the C3 activation
complex has a slightly affinity for C3b and C3b
binds to them.
 When C3b binds to C2b and C4b it forms a new
complex referred to as the C5 activation
complex.
THE C5 ACTIVATION COMPLEX

 The C5 activation complex (C2b,C4b,C3b)

activates C5 proteins by cleaving them into C5a
and C5b.
 Many C5b proteins are produced by the C5
activation complex. These C5b begin to coat
the surface of the bacteria.
THE FUNCTION OF C5A

 C5a disperses away from the bacteria.

- Binds to mast cells and increases inflammation.
- Most powerful chemotactic factor know for
leukocytes.
BUILDING THE MEMBRANE ATTACK COMPLEX

 C5b on the surface of bacteria binds to C6.

 The binding of C6 to C5b activates C6 so that it can
bind to C7.
 C7 binds to C8 which in turn binds to many C9’s.
 Together these proteins form a circular complex
called the Membrane attack complex (MAC).
 FORMATION OF C5B6789,
MEMBRANE-ATTACK COMPLEX
Cb5 attaches to C6, then to C7, and the
C5b67 inserts into the membrane.
↓
binding of C8 to membrane-bound
C5b67 induces a 10 Å pore.
↓
binding and polymerization of C9, a
perforin-like molecule, to C5b678.
↓
The completed membrane-attack
complex (MAC) has a tabular form and
functional pore size of 70 – 100 Å.
MEMBRANE ATTACK COMPLEX

 The MAC cause Cytolysis.

- The circular membrane
attack complex acts as a
channel in which
cytoplasm can rush out of
and water rushes in.
 The cells Inner integrity is
compromised and it dies.
 Animation of the classical
pathway.
THE ALTERNATIVE PATHWAY
 The alternative pathway is part of the non-specific
defense because it does not need antibodies to
initiate the pathway.
 The alternative pathway is slower than the Classical
pathway.
THE ALTERNATIVE PATHWAY IS AB-
INDEPENDENT

 The activation of alternative pathway doesn’t need Ab;

thus, It is a component of the innate immune system.
 It is initiated by cell-surface constituents that are
foreign to the host, e.g., bacterial cell wall.
 C1, C4 and C2 are not involved in the alternative
pathway.
 Four serum proteins, C3, factor B, factor D, and
properdin, are involved in this pathway.
INITIANTION OF THE ALTERNATIVE PATHWAY

 C3 contains in unstable
thioster bond.
 This unstable bond makes
C3 subject to slow
spontaneous hydrolysis to
C3b and C3a.
 The C3b is able to bind to
foreign surface antigens.
 Mammalian cells contain
sialic acid which inactivates
C3b.
FACTOR B

 C3b on the surface of a

foreign cells binds to
another plasma protein
called factor B.
FACTOR D

 The binding of C3b to

factor B allows a
protein enzyme called
Factor D to cleave
Factor B to Ba and Bb.
 Factor Bb remains
bound to C3b while Ba
and Factor D disperse
away.
THE C3 ACTIVATION COMPLEX

 Properdin, also called factor P, binds to the C3bBb

complex to stablize it.
 C3bBbP make up the C3 activation complex for the
alternative pathway.
THE C3 ACTIVATION COMPLEX
 The C3 activation
complex causes the
production of more
C3b.
 This allows the initial
steps of this pathway to
be repeated and
amplified.
 2X10₆ molecules can
be generated in 5
minutes.
C5 ACTIVATION COMPLEX

 When an additional
C3b binds to the C3
activation complex it
converts it into a C5
activation complex.
 The C5 activation
complex cleaves C5
into C5a and C5b.
 C5b begins the
production of the MAC.
THE LECTIN PATHWAY ORIGINATES WITH
HOST PROTEINS BINDING MICROBIAL
SURFACES

Lectin: Protein that bind to a carbohydrate.

MBL (mannose-binding lectin):

- an acute phase protein which binds to mannose residues
on glycoproteins or carbohydrates on the surface of
microorganisms (structurally similar to C1q)
MASP-1 & MASP-2 : MBL-associated serine protease
(structurally similar C1r and C1s)
- MBL is induced during inflammatory responses.
- After MBL binds to the surface of a microbe, MBL-
associated serine proteases, MASP-1 and MASP-2,
bind to MBL.
- The MBL-MASP-1/2 complex mimics the activity of
the C1r and C1s, and causes cleavage and
activation of C4 and C2.
- Thus, the lectin pathway is Ab-independent. It is an
important innate defense mechanism comparable
to the alternative pathway, but utilizing the
elements of the classical pathway, except for the
C1 proteins.
THE THREE COMPLEMENT PATHWAYS CONVERGE AT THE
MEMBRANE-ATTACK COMPLEX

The Three complement

pathways converge at
the production of an
active C5 convertase
↓
C5b6789
membrane-attack
complex (MAC)
REGULATION OF COMPLEMENT SYSTEM
1. C1 inhibitor :
 Important regulator of classic pathway
 A serine protease inhibitor (serpin)
 Irreversibly binds to amd inactivates C1r and C1s, as well as
MASP in lectin pathway

2. Factor H :
 Regulates alternative pathway
 Reduce amount of C5 convertase available
 With both cofactor activity for the factor I-mediated C3b
cleavage, and decay accelerating activity against C3bBb (C3
convertase)
3. Properdin :
 Protects C3b and stabilizes C3 convertase.

4. Factor :
 Cleaves cell-bound or fluid phase C3b and C4b
→ Inactivates C3b and C4b.

5. Decay accelerating factor (DAF) :

 Glycoprotein on surface of human cells.
 Prevents assembly of C3bBb or accelerates disassembly of
preformed convertase → no formation of MAC.
 Acts on both classical and alternative.
6. C4b-binding protein (C4BP) :
 Inhibits the action of C4b in classical pathway.
 Splits C4 convertase and is a cofactor for factor I.

7. Complement Receptor 1 (CR-1) :

 Co-factor for factor I, together with CD46

8. Protectin (CD59) and Vitronectin (S protein) :

 Inhibits formation of MAC by binding C5b678
 Presents on “self” cells to prevent complement from
damaging them
CLINICAL ASPECTS OF COMPLEMENT
1. Deficiency of C5-C8 & Mannan-binding lectin :
 Prodispose to serve Neisseria bacteremia.

2. Deficiency of C3 :
 Severe, recurrent pyogenic sinus & resp. tract infections.

3. Deficiency of C1 esterase inhibitor :

 Angiodema →inc. capillary permeability and edema.

4. Deficiency of DAF :
 Increased complement-mediated hemolysis → paroxysmal
nocturnal hemoglobinuria
5. Transfusion mismatches :
 Activation of complement → generate large amounts of
anaphylatoxins & MAC → red cell hemolysis.

6. Autoimmune diseases :
 Immune complexes bind complement → low complement
levels + activate inflammation → tissue damage.

7. Severe liver disease :

 Deficient complement proteins → predispose to infection
with pyogenic bacteria.
8. Factor I deficiency :
 Low levels of C3 in plasma due to unregulated activation of
alternative pathway → recurrent bacterial infections in
children.
 Mutation in factor I gene → implicated in development of
Hemolytic Uremic Syndrome.