The Complement System

Question 1

Functions of the complement system

Answer 1

Acascade of proteolytic cleavage
1. Lyse target cells
2. Opsonisation
3. Induction of inflammation by anaphylatoxins
4. Clear immune complexes
5. Virus neutralisation by aggregation, opsonisation, and lysis (of enveloped viruses) which reduces infectivity.

Question 2

Activation of the alternate pathway

Answer 2

Environment at the pathogen surface alters c3 conformation to resemble that of C3b

Question 3

Lectin pathway activation

Answer 3

Mannose-binding lectin binds the pathogen surface carbohydrates such as NAG to activate the cascade and C3b production

Question 4

Classical pathway activation

Answer 4

C-reactive protein or antibody binds antigens on the pathogen surface, activating the cascade….

Question 5

Unique feature of the alternative pathway

Answer 5

it does not rely on pathogen-binding and is instead initiated by hydrolysis of the C3

Question 6

First steps of the alternative pathway

Answer 6

1. “Tickover” - C3 undergoes spontaneous hydrolysis into C3(H2O) which binds to factor B, and then factor B is cleaved by Factor D, Bb remains attached - C3(H2O)Bb.
2. C3(H2O)Bb is a C3 convertase that cleaves more C3 into C3a/b. C3b is rapidly inactivated unless it binds the cell surface.

Question 7

Alternative pathway following binding of C3 to the cell surface

Answer 7

1. Factor B binds non covalently to the C3b and is cleaved by Facttor D into Bb - C3bBb.

Question 8

What is iC3b

Answer 8

C3b that has been bound to Factor H, CR1, and MCP is cleaved by Factor I to yield iC3b, the inactive form of C3b

Question 9

Factor P (AKA properdin) function

Answer 9

Binds to the C3bBb complex and stabilises to increase activity.

Question 10

How does C3b bind the cell surface

Answer 10

It binds covalently via the exposure of its highly reactive thioester bond. Hydrolysis of this bond is what causes inactivation and results in non-bound C3b.

Question 11

What is factor D

Answer 11

A serine protease that circulates in its active enzymatic form. It has no other structure than Factor B bound to C3b

Question 12

Factor H function

Answer 12

Binds C3b to compete with factor B and displace Bb from the C3 convertase, acting alongside factor I.
Only applies to the alternative pathway where Bb is involved.

Question 13

Factor I function

Answer 13

A plasma protease that cleaves C3b and C4b in the presence of cofactors such as Factor H, C4b-binding protein, and CD46.
This cleavage produces iCb

Question 14

DAF function

Answer 14

Also known as CD55
Competes with factor B for C3b binding and can displace Bb from already formed convertases

Question 15

MCP function

Answer 15

Also known as CD46
A host membrane protein

Question 16

CR1 function

Answer 16

Also known as CD35
A transmembrane glycoprotein receptor on cells that acts as a receptor for components C3b and C4b, displacing C2a and Bb respectively
Controls and attenuates the cascade and helps clear immune complexes.

Question 17

Protectin function

Answer 17

Also known as CD59
A host cell glycoprotein that binds complements C5b678, preventing the polymerisation of C9 and thus the formation of MAC.
These are not found on foreign cell surfaces.

Question 18

Sequence for the formation of the MAC

Answer 18

1. C5 is cleaved by a C5 convertase into C5b and C5a
2. C5b binds C6 and C7 before embedding in the cell membrane.
3. C8 binds the membrane-mound complex
4. Several molecules of C9 are recruited and polymerise to form a transmembrane pore.

Question 19

Anaphylatoxins

Answer 19

C3a and C5b increase blood vessel permeability allowing;
- complement and other plasma proteins to more readily enter infected tissues
- leukocytes to enter infected tissue and activate resident macrophages.

Question 20

MBL

Answer 20

Mannan-binding lectin binds to mannose residues and other sugars that are commonly accessible and arranged in a pattern that allows binding on pathogenic cells.
It does not bind vertebrate cells as these structures are covered by other sugar groups such as sialic acid.

Question 21

When is MBL produced

Answer 21

It exists in the plasma at low concentrations, but increases its production in the liver during the acute phase of the immune response - it is an acute phase protein.

Question 22

Steps of the lectin pathway following MBL binding to the cell

Answer 22

1. MASP-2 cleaves C4 into C4a and C4b. C4b attaches to the microbial surface.
2. MASP-2 cleaves C2 into C2a and C2b
3. C2a binds the C4b at the surface, producing the C3 convertase C4b2a.
4. C4b2a can then go on to cleave C3 molecules to produce C3b at the cell surface.

Question 23

What do soluble C3b molecule do other than bind directly to the cell surface?

Answer 23

They bind pre-existing C3 convertase at the cell surface to produce the appropriate C5 convertases.
- Classical/lectin - C3b joins C4b2a to form C4b2a3b
- Alternative - C3b joins C3bBb to form C3bBb3b

Question 24

C1 protein complex

Answer 24

C1q bound to two molecules each of C1s and C2s - C1q2s2r.
Binds to antibodies that have been complexed with antigens, as well as directly to the cell surface of a pathogen and undergoes autocatalysis.

Question 25

What happens when C1q binds an antigen

Answer 25

1. Causes a conformational change in the (C1r:C1s)2 complex, 2. leading to autocatalytic enzymatic activity in which C1r cleaves C1s into its active form as a serine protease. 3. C1s then cleaves C4. 4. C4b binds the cells surface as well as a molecule of C2 5. Bound C2 is more susceptible to cleavage, and the resulting C2b fragment remains bound to the C4b.

Question 26

What is C1s

Answer 26

A serine protease that has the ability to cleave C4 and C2 when cleaved by C1r

Question 27

What is C2b

Answer 27

A serine protease

Question 28

Homology of these three pathways

Answer 28

The MB-lectin and classical pathways use very similar proteins and both produce C4b2a. MASP-1 and MASP-2 are closely homologous to C1r and C1s, likely evolving from a gene duplication of a common precursor. The alternative pathway is thought to have evolved more recently.

Question 29

MBL deficiency

Answer 29

People with this condition experience more early childhood infections, illustrating the importance of innate host defences in childhood prior to the maturation of adaptive immune responses.

Question 30

Why dont classical and MBL pathways affect host cells (unrelated to inhibition by other molecules)

Answer 30

The enzymatic events of these events are confined to the same site so C3 activation only occurs at the pathogen surface and not in the plasma/host cells. This is achieved via the binding of the eactive thioester bond of C4b to the pathogen surface in its immediate vicinity. If it doesnt bind, the thioester bond is hydrolised which irreversibly inactivates C4b. Likewise, C2 is only susceptible to cleavage by C1s when bound to C4b and is thus confined to the pathogens surface. Ultimately, the only site opsonised is the pathogen.

Question 31

How is C5 convertase formed

Answer 31

C4b2a and C3Bb have the same activity in cleaving other C3 molecules into C3a and C3b. The C3b may either bind the cell surface or attach to the pre-existing C3 convertases to produce C5 convertases.

Question 32

Where do all three pathways converge?

Answer 32

At the formation of a C5 convertase, either in the form C4b2a3b or C3bBb3b.

Question 33

In what way are many complement components “labile”

Answer 33

The thioester bond on C4b and C3b that is exposed by cleavage of the original components is highly reactive and unless it binds to its targets, it is irreversibly hydrolised, cleaving the bond and inactivating the molecule.

Question 34

Where do the MBL and Classical pathways converge?

Answer 34

When C4b binds the pathogens surface, the cleavage of C2 is initiated via the respective proteases MASP-2 and C1s. Each of these produces C4b2a!!

Question 35

C5 convertase function

Answer 35

Captures C5 through binding on an acceptor site of 3b, making it susceptible to cleavage by the serine protease activity of C2a and C3b. This generates C5b and C5a.

Question 36

C5b function

Answer 36

Initiates the MAC by binding C6 and C7… exposing a site for C8 to bind to… exposing a site for C9…

Question 37

C1INH

Answer 37

Causes dissociation of C1, C1q separating from C1r2s2 A serine protease inhibitor that acts in the classical and lectin pathways.

Question 38

Factor I cofactor activity on C3b

Answer 38

With MCP, CR1, or Factor H, Factor I cleaves C3b into C3c, iC3b, and C3dg.

Question 39

Factor I cofactor activity on C4b

Answer 39

With MCP, CR1, or C4BP, Factor I cleaves C4b into C4c and C4d

Question 40

Cleavage of anaphylatoxins C3a and C5a

Answer 40

Each of carboxypeptidase N, carboxypeptidase B, and carboxypeptidase R cleaves the molecules into C3a des Arg and C5a des Arg respectively