Complement System: Activation & Functions

1. INTRODUCTION

Complement (C):

A system of factors that occur in normal serum.

Activated by Antigen–Antibody (An–Ab) interaction.

Subsequently mediates a number of biologically significant consequences.

It is a heat-labile component present in the normal sera of humans and animals.

📜 Historical Background

Buchner (1889): Discovered that the bactericidal effect of serum was destroyed by heating at 55°C for 1 hour → revealed its heat-labile nature.

The present name "Complement" was coined by Ehrlich, as this factor complemented the action of the antibody (Ab).

 

2. GENERAL PROPERTIES OF COMPLEMENT

Occurrence: Present in sera of all mammals and other animals (birds, amphibians, fish).

Non-specific serological reagent: Complement (C) from one species can react with antibodies from another species.

Efficiency of reaction is influenced by the taxonomic distance between the species.

Constitutes about 5% of normal serum protein.

Not increased by immunisation.

Heat labile as a whole, however, some components are heat stable.

Activity destroyed in 30 minutes at 56°C.

Does not bind to free antigen (An) or antibody (Ab) — only to Abs that are bound to their antigens.

Fixation / Binding / Consumption: 

Combining of 'C' with Ab is termed as fixation, binding, or consumption.

Leads to activation of the Classical Complement Pathway.

Immunoglobulin Classes that Fix Complement:

✅ Only IgM, IgG3, IgG1, and IgG2 (in this order) fix C.

❌ IgG4, IgA, IgD, and IgE do not fix complement.

Complement Binding Site:

Located on the Fc portion of Ab.

IgG → CH2 domain

IgM → CH4 domain

These domains are expressed only when Ab is bound to An.

Fixation of 'C' is not influenced by An nature, only by the class of Ig.

3. COMPONENTS OF COMPLEMENT SYSTEM

🧪 Complement System comprises at least 20 chemically and immunogenically distinct serum proteins, divided into:

Complement Components

Properdin System

Control Proteins

🔬 Complement Components(C)

A complex of nine different fractions → C1 to C9.

C1 occurs as a calcium ion–dependent complex with three subunits:

C1q, C1r, C1s

Thus, C is made up of 11 different proteins in total.

⚙️ Sequence of Cascade

C1 to C9 occur in sequence in a cascade reaction, except that C4 occurs between C1 and C2.

Correct order:

C1 → C4 → C2 → C3 → C5 → C6 → C7 → C8 → C9

📝 Nomenclature Rules

(a) Complement denoted as 'C'.

(b) Enzymatically activated components denoted with a bar → C1̄, C2̄, C3̄, etc.

(c) Fragments cleaved from C components during cascade indicated with small letters → C3a, C3b, etc.

(d) Inactivated forms denoted with prefix 'i' → iC3b, iC3a, etc.

4. COMPLEMENT ACTIVATION

Complement is usually present in the body in an inactive form.

Activity is induced by Ag–Ab combination or other stimuli.

C components then react in a specific sequence as a "cascade" (pass on to).

'C' – Cascade → A series of reactions:

Preceding components act as enzymes on the succeeding components and cleave them into dissimilar fragments:

1 Larger fragment → joins the cascade.

1 Smaller fragment → released; possesses biological effects. These fragments contribute to defence mechanisms by:

(a) Amplifying inflammatory processes

(b) Increasing vascular permeability

(c) Inducing smooth muscle contraction

(d) Chemotaxis of leucocytes

(e) Promoting virus neutralisation

(f) Detoxifying endotoxin

(g) Release of histamine from mast cells

Pathways of Complement Cascade:

C cascade is triggered off by two parallel but independent mechanisms/pathways and lectin pathway.

These pathways differ only in initial steps.

Once C3 is activated, subsequent steps are common to both pathways.

🔀 Three Pathways:

Classical Pathway: So called as it was the first one identified; represents specific acquired immunity.

Alternate (Properdin) Pathway: Represents a more primitive system of nonspecific innate immunity.

Lectin Pathway: The lectin pathway is similar to the classical pathway, but it does not require antibodies.

 I.  CLASSICAL PATHWAY OF COMPLEMENT ACTIVATION

In the Classical Pathway, C components react in a specific sequence following C1 activation. The pathway ends up in immune cytolysis.

🔹 Steps of the Classical Pathway

✅ STEP I – Recognition & C1 Activation

C1 antibody binds to the Ag–Ab complex.

The recognition site of C1 is C1q.

C1q reacts with the Fc portion of IgM or IgG.

C1q has 6 binding sites.

Effective activation occurs only if at least two binding sites of C1q are attached to Ig.

Hence, 1 molecule of IgM, or 2 molecules of IgG can induce the process.

C1q binding in the presence of calcium ions activates C1r and C1s.

✅ STEP II – C4 Cleavage by Activated C1s

Activated C1s acts as an esterase.

It cleaves several molecules of C4.

C4 splits into two fragments:

l C4a Acts as an anaphylatoxin

l C4b Binds to cell membranes along with C1

✅ STEP III – C2 Cleavage & Formation of C3 Convertase

C4b, in the presence of magnesium ions, cleaves C2.

C2 splits into two fragments:

l C2a Linked to cell-bound C4b

l C2b Released into the fluid phase

The complex C4b2a has enzymatic activity.

This complex is referred to as the Classical Pathway C3 Convertase.

✅ STEP IV – C3 Cleavage & Formation of C5 Convertase

C4b2a (C3 convertase) splits C3 into 2 fragments:

C3a Acts as an anaphylatoxin

C3b Bound along with C4b2a

C3b binds along with C4b2a to form a trimolecular complex: C4b2a3b

This trimolecular complex has enzymatic activity. It is called C5 Convertase.

✅ STEP V – Membrane Attack Phase of Complement

The membrane attack phase is the terminal phase of the complement cascade and results in the formation of the Membrane Attack Complex (MAC).

· C4b2a3b acts as the C5 convertase. 

· C5 convertase cleaves C5 into C5a and C5b.

· C5a acts as an anaphylatoxin involved in inflammation.

· C5b continues the complement cascade.

· C6 and C7 join C5b, forming a heat stable trimolecular complex, C5b67.

· A part of the C5b67 complex binds to the cell membrane and prepares it for lysis.

· C8 and C9 then join the complex, causing cell lysis.

· Unbound C5b67 can escape and amplify the reaction by adsorbing onto unsensitized “bystander cells,” making them susceptible to lysis by C8 and C9.

· C5b67 also has chemotactic activity. 

· The process results in complement-mediated cytolysis by producing holes approximately 100 Å in diameter in the cell membrane.

· These holes disrupt the osmotic integrity of the membrane.

· This leads to the release of cell contents and ultimately cell lysis.

· C5b, C6, C7, C8, and C9 form the Membrane Attack Complex (MAC). 

· The three complement pathways converge at the MAC.

Fig 1. Classical Pathway

Alternate Pathway of Complement Activation

· The central process in the complement cascade is C3 activation.

· In the classical pathway, C4b2a acts as the C3 convertase.

· Activation of C3 without the participation of C3 convertase is called the alternative pathway.

· The alternative pathway was first demonstrated by Pillemer (1954) as the properdin system.

· The properdin system consists of a group of about 30 serum proteins.

· These proteins contribute to antimicrobial defence without requiring specific antibodies.

· Zymosan, a polysaccharide from the yeast cell wall, can activate the properdin system.

· Other substances can also activate the pathway, such as bacterial endotoxins, IgA & D, and cobra venom factor.

· Hepatic factor (C3 factor) is also present in the serum of patients with glomerulonephritis.

✅ STEP I – Binding of C3b to an Activator

· C3b binds to an activator.

· C3b is continuously generated in small quantities in the circulation.

· In the free state, C3b is rapidly inactivated by serum factors H and I.

· Factors H and I are complement control proteins.

· When C3b is bound to an activator, it is protected from inactivation.

✅ STEP II- Formation of C3bB Complex

· Activated C3b interacts with the serum protein Factor B.

· Factor B is also called C3 proactivator. 

· C3b and Factor B form a calcium-dependent complex, C3bB.

✅ STEP III- Formation of C3 Convertase

· C3bB is cleaved by another serum protein called Factor D.

· Factor D is called the C3 proactivator convertase.

· Factor D cleaves C3bB into Ba and Bb.

· Ba is released into the medium.

· Bb remains bound to C3b.

· The resulting complex, C3bBb, is formed.

· C3bBb acts as the C3 convertase of the alternative pathway.

· The C3bBb complex is highly heat-labile.

· Properdin (Factor P) stabilizes the C3 convertase.

· The stabilized complex hydrolyzes C3 into C3a and C3b.

· The further steps are the same as in the classical pathway.

Fig 2. Alternate Pathway

II. Lectin pathway of complement

The lectin pathway is one of the 3 pathways of complement activation. It is antibody-independent and is activated when lectins recognize carbohydrates on the surface of microbes.

 

Mechanism

 

Microbial surface carbohydrates
↓
MBL (mannose-binding lectin) binds to mannose/N-acetylglucosamine on pathogen
↓
Activates MASP-1 and MASP-2 (MBL-associated serine proteases)
↓
C4 → C4a + C4b
C2 → C2a + C2b
↓
C4b2a = C3 convertase
↓
C3 → C3a + C3b
↓
C3b joins C4b2a → C4b2a3b = C5 convertase
↓
C5 → C5a + C5b
↓
C5b–C9 → Membrane Attak Complex (MAC)
↓
Cell lysis