BABS 2433 IMMUNOLOGY

Assignment 1
Name: Chong Khai En
Student ID: 15WAU08702
Group: RBS 2- Group 1
Question1:
What are the major difference between the alternative pathway and
the classical pathway?
The difference between the aternative pathway and the classical pathway is
their activation pathway. In alternative pathway, the activation occurs
independently of antibodies. It begins with the cleavage of C3 into C3a and C3b.
C3b then binds to microbial surfaces. On the other hand, the activation of
classical pathway is triggered by binding of IgM or certain subclasses of IgG bind
to antigens or microbial cell surface. Adjacent Fc regions of antibodies binds to
C1 component, and activated C1 will cleave two proteins C4 and C2.
The second difference is C3 convertase in alternative pathway and classical
pathway. In alternative pathway, the Bb fragment attaches to C3b, C3bBb
complex breaks down more C3 which functioning as C3 convertase in alternative
pathway. In classical pathway, C4b binds to C2 to form the C4bC2a complex
which is the C3 convertase of classical pathway.
In addition, the C5 convertase in both pathways also differs. In alternative
pathway, C3bBb molecules bind to an additional C3b molecule and the C3Bb3b
complex functions as a C5 convertase. In classical pathway, some of the C3b
binds to the C4b2a complex and form C4b2a3b complex which functions as a C5
convertase.
Furthermore, there are no additional proteins in classical pathway. In
alternative pathway, there are additional proteins such as factor B, factor D and
properdin. Factor D cleaves factor B and produce an enzyme composed of C3b
and Bb fragment. This enzyme is stabilized by properdin which cleaves more
molecules of C3 into C3a and C3b.
Alternative pathway

Classical pathway

The activation occurs independently of The activation is triggered by binding

antibodies. It begins with the cleavage of IgM or certain subclasses of IgG bind
of C3 into C3a and C3b. C3b then binds to antigens or microbial cell surface.
to microbial surfaces.
C3 convertase is C3bBb complex.

C3 convertase is C4bC2a complex.

C5 convertase is C3Bb3b complex.

C5 convertase is C4b2a3b complex.

Additional proteins such as factor B, No additional proteins.

factor D and properdin.

Question2:
Explain how the complement pathway can be regulated.
It is essential that activity be down-regulated on host cells while efficient
activation is allowed on foreign targets. Mammalian cells express regulatory
proteins to inhibit complement activation and thus preventing complementmediated damage to host cells.
Decay-accelerating factor (DAF) is a lipid-linked cell surface protein that can
disrupt the binding of Bb to C3b and the binding of C4b to C2a, thus blocking C3
convertase formation and terminating complement activation by both the
alternative and the classical pathways. DAF also decreases the stability of the
C3 convertase of the classic and lectin pathways, C4bC2a, by accelerating its
dissociation to C4b and C2a.
In addition, membrane cofactor protein (MCP) acts as a cofactor for the
proteolysis of C3b into inactive fragments, which mediated by a plasma enzyme
called factor I. CR1 may displace C3b and promote its degradation.
C1 inhibitor (C1 INH) is a regulatory protein that stops complement activation
early at the stage of C1 activation. C1 inhibitor irreversibly binds to and
inactivates C1r and C1s of the CP and MASP-1 and MASP-2 of the LP, thus
inhibiting the initiating steps of these activation pathways.
Furthermore, CD59 as known as protectin is the key regulator of the terminal
pathway. It inhibits C9 association with C5b-8 to prevent C5b-9 formation. Both
CD59 and DAF anchor to cell membranes by a glycosyl phosphatidylinositol
molecule.
Other additional regulatory proteins limit complement activation at the last
steps, for example MAC formation. Microbes lack the regulatory proteins and will
susceptible to complement. The regulation can be overwhelmed by too much
complement activation even in mammalian cells. Therefore, mammalian cells
can become targets of complement if they are coated with large amounts of
antibodies.
Inherited deficiencies of regulatory proteins cause uncontrolled and pathologic
complement activation. Deficiency of C1 INH is the cause of a disease called
hereditary angioneurotic edema. In this case, excessive C1 activation and the
production of vasoactive protein fragments lead to leakage of fluid (edema) in

the larynx and many other tissues. Paroxysmal nocturnal hemoglobinuria is a

disease results from the acquired deficiency in hematopoietic stem cells of an
enzyme that synthesized the glycolipid anchor for several cell surface proteins,
including the complement regulatory protein DAF and CD59. In these patients,
unregulated complement activation occurs on erythrocytes, leading to their
lysis. Deficiency of the regulatory protein factors H and I results in increased
complement activation and reduced levels of C3, causing increased
susceptibility to infection.

Question 3:
What are the differences between Mannose lectin complement pathway
and classical pathway?
The difference between Mannose lectin complement pathway and classical
pathway is their activation pathway. In classical pathway, the activation is
triggered by binding of IgM or certain subclasses of IgG to antigens. On the
other hand, the Mannose lectin complement pathway is not initiated by
antibodies but by the attachment of plasma mannose-binding lectin (MBL) to
microbes.
In order to active the complement system, MBL binds to another protein, a
serine protease called MASPs (MBL-associated serine proteases). When MBL
binds to its target, the MASP protein functions like a convertase to cleave C3
into C3a and C3b, and then activate C4. In other word, MBL is structurally
similar to a component of C1 of the classical pathway and functions to activate
C4. The subsequent steps are essentially the same as in the classical pathway.

Question 4:
For the process of inflammation, account for the following:
(a) the role of selectins, integrins and cohesive molecules in the
extravasation procedures.
Selectins are glycoproteins that are lectin, for example sugar-binding
molecules, some of which are expressed on leukocytes and some of endothelial
cells. Selectin is involved in the rolling process during extravasation. During
rolling process, E and P-selectin are produced by the endothelial cells on the
vessel walls of the circulatory system. These selectins can bind to sialyl-Lewis x
on the surface of the neutrophil and slowing down the neutrophil so that it rolls
along the endothelium. Furthermore, P-selectin also binds to P-selectin
glycoprotein ligand-1 (PSGL-1) which is modestly expressed on human

leukocytes. They also recruit lymphocytes to inflammation site, which are

indicative of the presence of pathogen. E and P selectin are important to our
bodys immune system because they help in acute immune responses by
directing lymphocytes to specific areas of infection in both innate and adaptive
immune systems.
Integrins are heterodimeric proteins consisting of - and -chains and they can
pair to result in many combinations of integrins with different expression and
binding specificity. Some integrins will bind to target molecules only following
activation of the leukocytes by various factors. Integrins are responsible for the
anchoring of cell to the ECM, cell to cell interactions and two-way signal
transmission, because they take part in pathways that carry signals both to and
from the cell. LFA-1 is lymphocyte function-associated antigen 1, can be found
on all T-cells and B cells, macrophages and neutrophils. During extravasotion,
integrin LFA-1 changes conformation and binds to ICAM-1 on antigen-presenting
cell, which functions as an adhesion molecule.

(b) What are the roles of cytokines and complement proteins during
inflammation.
Cytokines that involved during inflammation includes tumor necrosis factor-
(TNF), interleukin-1 (IL-1) and interleukin-6 (IL-6). TNF and IL-1 signal
endothelial cells causes them leaky to fluid, thereby leading to influx of plasma
which containing antibodies, complement components, etc. In addition, TNF
and IL-1 signal endothelial cells causes them sticky for leukocytes, and leading
to influx of neutrophils first, followed by monocytes and lymphocytes. On the
other hand, IL-6 promotes adaptive immune responses and has systemic effects.
Three of the cytokines are produced by macrophages which are TNF, IL-1 and
IL-8, they are important in the inflammatory response. TNF and IL-1 activate
endothelial cells lining the blood vessels at the infection site. This causes
endothelial cells to express on their cell surface molecules that neutrophils in
the blood stream can bind to. Thus, neutrophils can leave the bloodstream and
enter the tissue. Neutrophils and macrophages ingest and kill bacteria and other
microorganisms. IL-8 which is chemostatic for neutrophils also recruits
neutrophils.
Different plasma proteins make up the complement system which plays many
roles in resistance to infection. In a manner analogous to the clotting cascade, it
includes a series of pro-enzymes and related factors that sequentially activate
each other causing the production of many biologically active proteins. During

inflammatory responses, a complement component called C5a is produced that

increases vascular permeability. Moreover, C3a and C5a can cause mast-cell
degranulation, thereby amplifying the inflammatory process.