Proteins

Plasma

p l a s m a i s th e l i q u i d medi um of bl ood Separatlon eit plasma proteins

Th "
| (5 5 -6 0 % ),i n w h i c h th e c e l l components-
The total concentrationof plasma proteins is
namely erythrocytes, leukocytes, platelets-are
about 6-8 g/dl. The plasma is a complex mixture
suspended.lf blood containing anticoagulants
(e.g. heparin, potassiumoxalate) is centrifuged, of proteins,and severaltechniquesare employed
to separatethem. An age-oldtechnique is based
the plasma separatesout as a supernatantwhile
on the use of varying concentrations of
the cells remain at the bottom. The packed cell
ammoni um sul fate or sodi um sul fate. By t his
volume or hematocrit is about 45%.
method, which is known as salting out process,
T h e te rm s e ru m i s a p p l i e d to the l i qui d the plasma proteins can be separated into
medium which separates out afterthe blood clots three groups-namel y al bumi n, gl obul ins and
(coagulates). Serum does not contain fibrinogen fi bri nogen.
and other clotting factors. Thus, the main
difference between plasma and serum is the Electrophoresis: This is the most commonly
presence or absence of fibrinogen. employedanalyticaltechniquefor the separation
of plasma (serum)proteins.The basic principles
lFt's6l{,.ti,rrr*+ot $f ood of electrophoresisare described in Chapter 43.
T h e to ta l v o l u me o f b l o o d i n an adul t i s Paper or agar gel electrophoresiswith vernol
around 4.5 to 5 liters. Blood performs several buffer (pH-8.6) separatesplasma proteins into
diversifiedfunctions. These include respiration, 5 distinct bands namely alhumin, a1, a2, B and
excretion, acid-base maintenance, water y globulins (Fig.9.l). The concentration of each
balance,transportof metabolites,hormonesand one of these fractions can be estimated by a
drugs, body defenseand coagulation. densitometer.

182
Chapter 9: PLASMAPROTEINS 183

Stan 2. E xcreti onof al bumi n i nto uri ne i n ki dney

damage.
3. Increased production of globulins
associated with chronic infections, multiple
myelomasetc.

Gomponents of plasma proteins

Globulins
The important plasma proteins along with
their characteristics(based on electrophoretic
pattern) and major functions are given in
Table 9.1. Some selected plasma proteins are
discussedhereunder.

Albumin is the major constituent (600/o)ol

of plasma proteins-
Fig.9.1 : Electrophoresis plasma proteinswith a concentrationof 3.5-5.0
,i::,mliiftffi lffitl-ii{F ,tH 'j ' g/dl. Human albumin has a molecularweight of
69,000, and consists of a single polypeptide
chai n of 585 ami no aci ds w i th 17 di sul fi de
Abnormal electrophoretic pattern
bonds.
Electrophoresis of serum proteins is
conveniently used for the diagnosisof certain Synthesis of albumin
diseases A l bumi n i s excl usi vel ysynthesi zedby the
1. Multiple myeloma : A sharp and distinct liver. For this reason, measurementof serum
M band appearsin the lglobulin fraction. albumin concentrationis conveniently used to
2. Acute infections : c1- and a2- globulins assessIiver function (synthesis decreasedin liver
are increased. diseases).Liver produces about 12 g albumin per
day which represents25'/. of the total hepatic
3. Nephrotic syndrome : Decreasedalbumin
protein synthesis.Albumin has an half-lifeof 20
with sharp and prominent c,2-globulin.
days.
4. Primary immune deficiency : Diminished
y globulin ba n d . Functions of albumin
5. a1-Antitrypsindeficiency: Diminishedcr1- Plasmaalbumin performsosmotic, transport
globulin ban d .
and nutritive functions
Albumin/globulin (A/G) ratio : The albumin 1. Osmotic function : Due to its high
concentrationof plasma is 3.5 to 5.0 g/dl while concentration and low molecular weight,
that of total globulins is 2.5 to 3.5 g/dl. The albumin contributes to 75-8oo/o of the total
normal A/G ratio is 1.2 to 1.5 : 1. The A,/C ratio plasma osmotic pressure (25 mm Hg). Thus,
is lowered either due to decreasein albumin or al bumi n pl aysa predomi nantrol e i n mai ntai ni ng
increasein globulins, as found in the following blood volume and body fluid distribution.
c ondit ions Decreasein plasma albumin level results in a
1. Decreasedsynthesisof albumin by liver- fall in osmotic pressure,leading to enhanced
usuallyfound in liver diseasesand severeprotein fluid retention in tissuespaces/causingedema.
m alnut r it ion . The edema observed in l<washiorkor,a disorder
184 BIOCHEMISTF|Y

Plasma Molecular Maior function(s)

concentration weight

Albumin 3.5-5.0g/dl nutritive

transport,
69,000 Osmotic, andbuffering
Prealbumin 2$-30mg/dl 61,000 Transports to someextent
thyroxine
a,-Globulins 0.3.{,5 g/dl
c,-Antitrypsin < 0.2 g/dl 54,000 Inhibitor
of trypsin
(HDL)
c[1-Lipoproteins 0.2{.3 g/dl Transports andphospholipids
cholesterol
Orosomucoid < 0.1g/dl 44,000 Bindswithprogesterone
Retinol protein
binding (RBP) 3-6 mg/dl 21,000 Transports A
vitamin
Thyroxine globulin
binding (TBG) 1-2 mg/dl 58,000 Transportsthyroidhormones
Transcortin
or cortisol 3-4 mg/dl of steroid
52,000 Majortransporter (e.9.
hormones
protein
binding (CBG) cortisol,
corticosterone)
oq-Globulins 0.4-0.8g/dl
o9-Macroglobulin 0.2{.3 g/dl andantiplasmin
800,000 Antitrypsin activity
Haptoglobins < 0.3g/dl withplasma
90,000 Binds and
freehemoglobin
(Hp1-1;Hp2-1andHp2-21 prevents
itsexcretion
Prothrombin < 0.02g/dl 63,000 Participates
in bloodcoagulation
Ceruloplasmin < 0.03g/dl 150,000 Transport
of copper; of Fe2*to Fe$.
oxidation
p-Globullns g/dl
0.6-1.1
p-Lipoproteins
(LDL) 0.2{.5 g/dl Transports
triacylglycerols
andcholesterol
Transfenin 0.24.3g/dl 76,000 Transports
iron
Hemopexin < 0.1g/dl 57,000 Transports
heme
Plasminogen < 0.05g/dl plasmin,
140,000 Forms involved
in fibrinolysis
yGlobullns 0.8-1.8mg/dl functions
Antibody
(fmmunoglobulins-lgG,
lgA,lgM,lgDandlgE;relerTable
9.2lordetails)
Fibdnogen 0.2-0.4g/dl in bloodcoagulation
340,000 Participates

of protein-energymalnutrition, is attributed to a prealbumin, retinol binding protein, thyroxine

drastic reduction in plasma albumin level. binding protein,transcortinand others as stated
in the functionsof plasmaproteinsin Table9.11.
2. Transport functions : Plasma albumin
binds to several biochemically important 3. Nutritive functions : Albumin servesas a
compounds and transports them in the sourceof amino acidsfor tissueprotein synthesis
circulation. These include free fatty acids, to a limited extent, particularly in nutritional
bilirubin, steroidhormones,calcium and copper. deprivationof amino acids.

[Note : Besidesalbumin, there are several 4. Buffering function : Among the plasma
other plasma transport proteins. These include proteins, albumin has the maximum buffering
f ira!"r!;*s"S : PLASMA PFIOTEINS 18s

capacity. However, the buffering action of Emphysema(Greek: emphusan-to inflate) is

a lbum in in plas m ai s n o t s i g n i fi c a nct o mp a re dto a term usedto representthe abnormaldistension
bicarbonatebuffer system. of lungs by air. At least5% of emphysemacases
are due to the deficiency of a1-AT. This is
,'t ; I ::;,-:,! $ Ea;,{,fi
I f *{: en{:t'* *,}{ o*! *eet:'i:i;,,;
; associatedwith lung infections(e.g.pneumonia)
1 . A lbum in, b i n d i n gto c e rta i nc o m p o u n d si n and increasein the activity of macrophagesto
the plasma, prevents them from crossing the releaseelastasethat damageslung tissues.In the
b l ood- br ain ba rri e r e .B . a l b u m i n -b i l i ru b i n normal circumstances, elastase activity is
complex, albumin-freefatty acid complex. i nhi bi tedby a1-A T.

2. Hypoalbuminemia (lowered plasma Effectof smoking on crl-AT : The amino acid

a lbum in) is obs e rv e di n ma l n u tri ti o n ,n e p h ro t i c methi oni neat posi ti on358 of a1-A T i s i nvol ved
sv ndr om eand c i rrh o s i so f l i v e r. i n bi ndi ng w i th proteases.S moki ng causes
oxi dati on of thi s methi oni ne to methi oni ne
3. A lbum in is e x c re te d i n to u ri n e
sulfoxide. As a result, a1-AT with methionine
(albuminuria) in nephrotic syndrome and in
sulfoxide cannot bind and inactivate proteases.
certain inflammatoryconditionsof urinary tract.
Emphysemais more commonly associatedwith
Microalbuminuria (3O-3O0mg/day) is cl i n ically
heavy smokingand the situationbecomesworse
important for predictingthe future risk of renal
in personswith cr1-AT deficiency.
diseases(Refer Chapter 36).
cl1-Antitrypsindeficiency and liver disease :
4. A lbum in is th e ra p e u ti c a l l yu s e fu l fo r th e
Thi s i s due to the accumul ati onof a mutant
treatmentof burns and hemorrhage.
o1-AT which aggregatesto form polymers.These
polymers,in turn-by an unknown mechanism-
cause liver damage (hepatitis) followed by
accumul ati onof col l agen resul ti ng i n fi brosi s
Globulins constituteseveralproteinsthat are (ci rrhosi s).
separatedinto four distinct bands (cr1,42, p and
gr
1-globulins)on electrophoresis(See Fig.9.l). ix ]-11#flc FsffiBrok$Eii
C lobulins , in g e n e ra l , a re b i g g e r i n s i ze It i s a hi gh mol ecul ar w ei ght (8,00,000)
th an album in. T h e y p e rfo rm a v a ri e ty o f protein and is a major constituentof u,2-fraction.
fu nc t ionswhic h in c l u d etra n s p o rta n d i m m u n i ty. cr2-Macroglobulininhibits proteaseactivity and
ln Table 9.1, the importantglobulins are given, servesas an anticoagulant.lts concentrationin
some of them are discussedhereunder. plasma is elevated in nephrotic syndrome. This
r t . 3 ' 1 } 5 { Y ,r
is due to the fact that majority of the low
mol ecul ar w ei ght protei ns are l ost i n uri ne
u1-Antitrypsin,more recentlycalled as a-anti- (proteinuria)in this disorder.
proteinase, is a glycoproteinwith 394 amino acids
a,rd a molecularweight of 54,000. lt is a major H A P TOGLOB IN
constituent of u1-globulin fraction of plasma
H aptogl obi n (H p) i s a pl asma gl ycoprotei n
:"oteinswith a normalconcentrationof about200
-q, dl. crl-Antitrypsinis a serineproteaseinhibitor. with an approximate molecular weight of
90,000. Hp is an acute phaseprotein since its
: combines with trypsin, elastase and other
plasma concentration is increased in several
:"o:easeenzymesand inhibitstheir activity.
i nflammatoryconditions.
' - :,i r i:11.-.::;:-n{:t+

' i " v t * 5 U *t,' * *;c,!yle;;td'i1'rt i ge

$ a;S !'*,4p'i r;e6 d# A;r

a. - { nt it r y ps ind e fi c i e n c yh a sb e e n i mp l i c a te d Haptoglobin binds with the free hemoglobin

- :,ro diseases,namely, emphysemaand a1-AT (known as extra-corpuscularhemoglobin) that
&ficiengy liver disease. spi l l s i nto the pl asma due to hemol ysi s.The
 d
186 B IOC H E MIS TRY

haptoglobi n-hemoglobin (Hp-Hb) complex (mol.

wt. 155,000) cannot pass through glomeruli of
k id n e y w h i l e fre e H b (m o l . w t. 65,000) can.
o)
Haptoglobin, therefore, prevents the loss of free E
Hb i n to u ri n e . IE
o
Clinical significance of Hp : Hemolytic E
anemia is associatedwith decreased plasma
a
concentrationof haptoglobin.This is explained
as follows. The haltt-life of Hp is about 5 days
wh i l e th a t o f H p -H b c o mp l e x i s 90 mi n. l n 45678910fi12
hemolytic anemia,free Hb in plasma is elevated Days ----*
leading to increased formation of Hp-Hb
Fig.9.2 : The responseof C-reactiveprotein (CRP) in
c om p l e x . T h i s c o mp l e x , i n turn, i s rapi dl y response to surgery (The normal acute Phase is depicted
cleared from the plasma resulting in decreased by blue line, the development ot infection by red line and
Hp l e v e l s . the response after treatment by green line).

CERULOPLASMIN
proteins decreases,and they are regarded as
Ceruloplasminis a blue colou;'ed,copper- negative acute phase reactants e.g. albumin,
c on ta i n i n ga 2 -g l o b u l i nw i th a m o l ecul arw ei ght transferrin.
of 150,000.lts plasmaconcentrationis about 30
mg/dl. Ceruloplasminbinds with almost 9O"/"oI C-reactive Frotein (CRPI
plasma copper (6 atoms of Cu bind to a
CRP is a maior component of acute phase
m o l e c u l e ).T h i s b i n d i n g i s ra th e rti ght and, as a
proteins.lt is producedin the liver and is present
result,copper from ceruloplasminis not readily
in the circulation in minute concentration
releasedto the tissues.Albumin carrying only
(< 1 mgidl). C-reactive protein (C strands for
1)oh of plasma copper is the major supplier of
carbohydrateto which it binds on the capsuleof
copper to the tissues.Ceruloplasminpossesses
pneumococi) is involved in the promotion of
oxidase activity, and it is associated with
immune system through the activation of
Wilson's disease which is discussed under
complement cascade.
copper metabolism (Chapter 1A.
Estimationof CRP in serum is important for
TRANSFERRIN the evaluation of acute phase response.The
response of CRP to surgery is depicted in
Transferrin (T0 is a glycoprotein with a
Fig.9.2. ln a normal surgery, serum CRP
molecularweight of 76,000. lt is associatedwith
p-globulin fraction.Tf is a transporterof iron in increasesand returns to normal level within
7-10 days. lf the recoveryis complicatedby any
t he c i rc u l a ti o n .
infection, it will be reflectedby the continuous
elevation of CRP which requires further
ACUTE PI{ASE PROTEINS
treatment.
Acute phaseresponserefersto a non-specific
responseto the stimulus of infection, injury,
variousinflammatoryconditions(affectingtissue/
organs),canceretc. This phaseis associatedwith
a characteristicpattern of changes in certain
plasmaproteins,collectivelyreferredto as acute The higher vertebrates,including man, have
phaseproteinse.g. o,1-antitrypsin, ceruloplasmin, evolved a defense system to protect themselves
complementproteins,C-reactiveprotein. During against the invasion of foreign substances-a
the acute phase, synthesisof certain plasma virus, a bacterium or a protein. The defense
Chapter 9 : PLASMA PFIOTEINS 187

lnterchain
disulfidebonds
*Hr N
rN H s
t-"'=\

*iirN 1-

F-', NH-r

Fab

S -S
Hingeregion
S_

cHo

cH2
Fc
Intrachain
disulfide
bonds

coo-

of the body are collectively referredto

strategies chains(mol. wt. 53,000 to 75,000 each)and two
as immunity, and are briefly described under identical light (L\ chains(mol. wt. 23,000 each)
immunology (Chapter42). lmmunoglobulins(or hel d together by di sul fi de l i nkagesand non-
antibodies)are describedhere. covalent interactions(Fi9,9.3\.Thus, immuno-
globulin is a Y-shaped tetramer (HzLz). Each
lmrmunoglobulins-basic concepts heavy chain contains approximately450 amino
lm m unoglo b u l i n s ,a s p e c i a l i s e d g ro u p of acids while each light chain has 212 amino
proteins are mostly associated with y-globulin acids. The heavy chains of Ig are linked to
fraction (on electrophoresis) of plasma proteins. carbohvdrates, hence i mmunogl obul i ns are
Some immunoglobulinshowever,separatealong glycoproteins.
with P and a-globulins.Therefore,it should be Constant and variable regions : Each chain
not ed t hat y-g l o b u l i n a n d i m m u n o g l o bul i n (L or H) of lg has two regions(domains),namely
are not synonymous. lmmunoglobulin is a the constant and the variable. The amino
functional ferm while y-globulin is a physical termi nal hal f of the l i ght chai n i s the vari abl e
term. region (V1)while the carboxyterminal half is the
constant region (Cg). As regards heavy chain,
S t r uc t ur e o f i s n m u n o g l o b u l i n s approximatelyone-quarterof the amino terminal
All the immunog:lobulin (Ig) molecules region is variable(Vx) while the remainingthree-
oasigally consist of two identical heavy (H) quartersis constant(Cs,, Csr, Csr). The amino
188 BIOCHEMISTFIY

Type H-Chain L{hains Molecular Molecular Percentage Serumconc, Itaior fundion(s)

formula werght carbohydrate mC/dl

IgG rorl, y2r2or y2)r2 -150,000 80f1,500 Mostly

responsible
for
immunity
humoral
IgA rorl, (o2rj1aor -(160,000h-i 15G.400 Protects
the body
(oraldr+ surfaces

IgM rorl, p2rj5 or - 900,000 12 50-200 Humoralimmunity,

ItzT"zls serves
asfirstlineof
defense
IgD rorl, (Qr2or Qt2) -180,000 13 1-10 B-cell
receptor?
IgE rorl, E2K2Ol e2?u2 -190,000 12 0.02-0.05 Humoralsensitivity
andhistamine
release,

acid sequence (with its tertiary structure) of l mmunogl obul i n G (IgGl

variable regions of light and heavy chains is
IgC is the most abundant (75-80%) class of
responsible for the specific binding of
i mmunogl obul i ns.
IgC i s composedof a single
im m u n o g l o b u l i n(a n ti b o d y )w i th anti gen.
Y-shapedunit (monomer).lt can traverseblood
Proteolytic cleavage of Ig : An immuno- vesselsreadily.IgG is the only immunoglobulin
globulin can be split by the enzyme papain to that can cross the placenta and transfer the
their fragments.These are two identical antigen mother's immunity to the developingfetus.IgG
binding fragments(Fab) and one crystallizable triggers foreign cell destruction mediated by
fragment(Fc).Papaincleavesthe immunoglobin complement system.
molecule at the site between Cp,1 and Cp2
regions which is referred to as hinge region. Innmunogl obul i n A { fgA }

IgA occurs as a single (monomer)or double

CLASSESOF IMMUNOGLOBULINS unit (dimer)held togetherby J chain. lt is mostly
Humans have five classes of immuno- found in the body secretionssuch as saliva,tears,
globufins-namely IgG, IgA, IgM, IgD and sweat,milk and the walls of intestine.IgA is the
IgE-containing the heavy chainsy, c, p, 6 and most predominantantibody in the colostrum,the
E, respectively. The type of heavy chain initial secretion from the mother's breast after a
ultimatelydeterminesthe classand the function baby is born. The IgA molecules bind with
of a given lg. bacterial antigens present on the body (outer
epithelial) surfaces and remove them. In this
Two types of light chains-namely kappa (r) way, IgA prcvents the foreign substances from
and lambda (l.)-are found in immunoglobulins. enteringthe body cells.
They differ in their structurein C1 regions.An
immunoglobulin(of any class)containstwo K or l rnmunogl obul i n M (IgM!
two l, light chains and never a mixture. The
l gM i s the l argesti mmunogl obul i ncomp osed
oc c u rre n c e o f r c h a i n s i s m o r e common i n
of 5 Y-shapedunits (IgC type) held togetherby
hum a n i mmu n o g l o b u l i n sth a n l , chai ns.
a J polypeptidechain. Thus IgM is a pentamer.
The characteristics of the 5 classesof human Due to its large size, IgM cannot traverseblood
immunoglobulinsare given in Table 9.2. vessels,hence it is restrictedto the blood stream.
Ghapter 9 : PLASMA PROTEINS 189

IgM is the first antibody to be produced in population. Femalesare more susceptiblethan

responseto an antigen and is the most effective males for this disorder and it usually occurs in
against invading microorganisms. lt may be the age group 45-60 years.
notedthat IgM can simultaneouslycombine with Abnormal lg production : Multiple myeloma is
5 antigenicsitesdue to its pentamericstructure. due to the malignancyof a singleclone of plasma
D (Ig D ! cells in the bone marrow. This results in the
I m m unog l o b u l i n
overproduction of abnormal immunoglobulins,
IgD is composed of a single Y-shaped unit mostly (75%) IgG and in some cases(25"h) IgA
and is present in a low concentration in the or IgM. IgD type multiple myeloma found in
circulation. IgD molecules are present on the younger adults is less common (<2%) but more
surfaceof B cells.Their function, however,is not severe. In patients of multiple myeloma, the
known for certain. Some workers believe that synthesi s of normal i mmunogl obul i ns i s
IgD may function as B-cell receptor. diminishedcausing depressedimmunity. Hence
recurrentinfectionsare common in thesepatients.
lmmunoglobulin E (IgEl
Electrophoretic pattern : The plasma of
IgE is a singleY-shapedmonomer.lt is normally
multiple myelomapatientsshowsa characteristic
presentin minuteconcentrationin blood. IgE levels
pattern of electrophoresis.
There is a sharp and
are elevated in individuals with allergiesas it is
distinct band (M band, for myeloma globulin)
associatedwith the body's allergic responses.The
between p-and y-globulins.Further,this M band
IgE moleculestightly bind with mast cells which
almost replacesthe y-globulin band due to the
releasehistamineand causeallergy,
diminished synthesisof normal y-globulins.
Production of ;mmunoglobulins Bencefones proteins : Henry BenceJonesfirst
by m ult ip l e Ee n e s described them in 1847. These are the light
(r or l,) of immunoglobulins that are
A s alr e a d y d i s c u s s e d ,i mmu n o g l o b ul i nsare chains
composedof light and heavy chains. Each light synthesized in excess.Bence Jonesproteins have
chain is produced by 3 separategenes,namely a molecular weight of 20,000 or 40,000 (for
a variable region (V1) gene/ a constant region dimer). In about 2O"/oof the patientsof multiple
(Cl) gene and a joining region (J) gene. Each myeloma, Bence .fonesproteins are excreted in
heavy chain is produced by at least 4 different urine which often damagesthe renal tubules.
genes-a variable region (Vg) gene, a constant Amyloidosis is characterized by the deposits
region (Cg) gene, a joining region U) gene and of light chain fragments in the tissue (liver,
diversity region (D) gene. Thus multiple genes kidney, intestine)of multiple myeloma patients.
are responsiblefor the synthesisof any one of
The presenceof BenceJonesproteinsin urine
t he im m u n o g l o b u l i n s .
can be detected by specific tests.
Antibody diversity : A person is capable of
1 . Electrophoresis of a concentrated urine is
generating antibodies to almost an unlimited
the best test to detect Bence Jones proteins in
range of antigens (more than one billion!). lt
ufl ne.
should, however, be rememberedthat humans
do not contain millions of genes to separately 2. The classical heat test involves the
c ode f or i n d i v i d u a li mmu n o g l o b u l i nmol ecul es. precipitation of Bence Jones proteins when
The antibodydiversityis achievedby two special slightly acidified urine is heatedto 40-50'C. This
processes/ namely comhination of various precipitate redissolves on further heating of urine
structural genes and somatic mutations, to boiling point. lt reappears again on cooling
urine to about 70oC.
MULTIPLE MYELOMA
3. Bradshaw's test involves layering of urine
Multiple myeloma, a plasma cell cancer, on concentratedHCI that forms a white ring of
constitutesabout\ 1'/" of all cancers affecting the precipitate, if Bence Jones proteins are present.
190 ., B IOC H E MIS TR Y

lntrinsic Extrinsic
patnway parhway
I
{z I
FactorX

Prothrombin
t,lftThro bin(ra)

Fibrin
Fibrinogen(l) (bloodclot)

Fig.9.4: Overuiew of blood cloning with the final common pathway-

two of these factors are designatedby a Roman

numeral. lt should, however, be noted that the
numbers representthe order of their discovery
The term hemostasis is applied to the
and not the order of their action. The cascadeof
sequence of physiological responsesto stop
blood clotting process is depicted in Fig.9.5 and
bleeding (loss of blood after an injury). This is
the salient features are discussedbelow. The
carried out by blood clotting.
active form of a factor is designated by a
Blood clotting or coagulation is the body's subscript a. The active clotting factors (with
major defense mechanism against blood loss. A exception of fibrin) are serine proteases.
blood clot is formed as a result of a seriesof
reactions involving nearly 20 different Sonversion of librinogen to fibrin
substances,most of them being glycoproteins, Fibrinogen(factor l) is a soluble glycoprotein
synthesized by the liver. that constitutes2-3"/"of plasmaproteins(plasma
Blood clotting process involves two concentration0.3 g/dl). Fibrinogenconsistsof 6
independentpathways polypeptide chains-two A cr, two B p and two 1
making the structure(A ct)z (B F)z'lz.
1. The extrinsicpathway is the initial process
in clotting and involvesthe factorsthat are not Fibrinogen undergoes proteolytic cleavage
presentin the blood (hence the name). catalysed by thrombin to release small
fibrinopeptides (A and B), This results in the
2. The intrinsic pathway involves a seriesof
formation of fibrin monomers which can stick
reactions participated by the factors present in
together to form hard clots (Fig.9.6). Clot
the blood.
formation is further stabilizedby covalentcross-
Strictly speaking,the extrinsic and intrinsic l i nki ng betw eengl utami neand l ysi ne resi dues.
pathways are not independent,since they are Thi s reacti on cross-l i nksfi bri n cl ots and is
coupled together.Further,the final reactionsare catalysedby fibrin stabilizingfactor (Xlll). The
identical for both pathways that ultimately lead red colour of the clot is due to the presenceof
to the activationof prothrombinto thrombin and red cells entangledin the fibrin cross-links.
the conversion of fibrinogen to fibrin clot
lFig.e.4). Conversion of prothromhin
to thronnbin
The blood coagulation factors in human
plasma along with their common names and Prothrombin(ll) is the inactivezymogenform
molecularweightsare listed in Table9"3. All but of thrombin (lla). The activationof prothrombin
Chapter I : PLASMA PROTEINS 191

occurson the plateletsand requiresthe presence (lX). The Christmasfactor is also activated by
of factorsVa and Xa, besidesphospholipidsand active proconvertin(Vlla).
Ca2+ . In the next step, the Staurt factor (X) is
activated by Christmas factor (lXa) and this
The extrinsic pathwas* reactionrequiresthe presenceof antihemophilic
The extrinsic pathway is very rapid and factor (Vllla), Ca2+and phospholipids.
occurs in responseto fissueinjury. This pathway The extrinsic and intrinsic pathways lead to
essentiallv involves the conversion of the formation of factor Xa which then
proconvertin(Vll) to its activeform (Vlla)and the participates in the final common pathway to
generation factor Xa. The tissue factor (lll), ultimately result in the formation of fibrin clot.
found to be necessaryto accelerate the action
Vlla on a factor X, is presentin lung and brain. Anticoagulants
Severalsubstances,known as anticoagulants,
T he int r ins ic p a th w a y
are i n use to i nhi bi t the bl ood cl otti ng.C al ci um
The intrinsic pathway is rather slow. lt is essentiallyrequired for certain reactions of
involves the participation of a contact system blood coagulation.The substanceswhich bind
(wounded surface) and a series of factors to with Ca2+ are very effective as anticoagulants.
generate factor Xa. These include oxalate, fluoride, EDTA and
citrate.
The Hagemanfactor (Xll) is activated(Xlla)on
exposureto activatingwound surfacecontaining Heparin is an anticoagulantused to maintain
collagen or platelet membranes.The formation normal hemostasis.lt is a heteropolysaccharide
of Xlla is acceleratedby kallikrein and HMK. found i n many ti ssuesi ncl udi ngmastcel l s i n the
The activated Hageman factor (Xlla) activates endotheliumof blood vessels.Heparincombines
factor Xl. The Xla activate the Christmas factor w i th anti thrombi nl l l w hi ch i n turn. i nhi bi tsthe

Factor number Common name(s) Subunit molecular weight

I Fibrinogen 340,000
tl Prothrombin 720,000
ill Tissuefactor,
thromboplastin 370,000
IV Calcium (Ca2*)
V labilelactor
Proaccelerin, 330,000
vtl Proconvedin,serumprothrombin conversion (SPCA)
accelerator 50,000
vill Antihemophilicfactor globulin
A, antihemophilic (AHG) 330,000
IX Christmasfactor, factorB,
antihemophilic 56,000
Plasma thromboplastincomponent (PTC)
X Staurt-Prowerfactor 56,000
XI Plasma thromboplastinantecedent (PTA) 160,000
xtl Hageman factor 80,000
xill factor(FSF),
Fibrin-stabilizing LikiLorand
fibrinoligase, factor 320,000
Prekallikrein 88,000
Highmolecular weightkininogen(HMK) 150,000
Note: Thenunbersrepresenttheoderof theirdisnveryaN nottheorderof thehaction.FactorVawason@refenedto as factorVl,
hencethereis nQfactorVl.
192 BIOCHEMISTFIY

Prekallikrein

FactorXll

Ertrinsic i
FactorXl pathway i

FactorlX Vl la+-,------: Factor Vl I i

FactorVl||----;*-=-+ VlI Factorlll

FactorX

lr ilr ' li:!.) n Thrombin(lla)

|J;Ll.liilrJV

Fibrinogen(l)

ractoJxttt---.------+ x tt
Fibrin
(hardclot)

Fig.9,5 : The blood clotting cascade in humans

(the active forms of the factors are reprcsented in red with subscript'a').

clotting factors ll, lX, X, Xl, Xll and kallikrein. Streptokinase is a therapeutic fibrinolytic
Heparin can be administeredto patientsduring agentwhich activates pl asmi nogen.
and after surgery to retard blood clotting.
The blood contains another anticoagulant-
namely protein C-which is activated by
thrombin. Active protein C hydrolyses and
inactivatesclotting factorsV and Vlll. FlMnogen
II
Warfarin, a vitamin K antagonist may be ThrombinI
consideredas an oral anticoagulanf. This acts by l9 Fibrinope A andB
reducingthe synthesisof certain clotting factors Y
Itr
( ll, Vl l , l X a n d X). \_./------\_/-------\_-/
Fibrinmonorner
i";nlEr"ls*olneii*,
Y
The term fibrinolysisrefersto the dissolution
or lysis of blood clots. Plasmin is mostly
responsiblefor the dissolution of fibrin clots.
Plasminogen,synthesizedin the kidney, is the Flbrinclot
inactive precursor of plasmin. Tissue
plasminogen activator (TPA) and urokinase Flg.9.6 : Diagrammatic representation
of
fibrinclot formationfromfibrinogen.
convert plasminogento plasmin.
frrr:r:,tslr ii : PLASMA PBOTEINS 193

i :\.l 4 f1 +." +:ri i i l+.l r t' , i i.r ir :::i' 1: :' ,r .:i' :' and sufferfrom internalbleeding (particularlyin
j oi nts and gastroi ntesti nal
tract). H emophi l i aA
Severalabnormalitiesassociatedwith blood
has gained importancedue to the fact that the
clotting are known. These are due to defectsin
Royal familiesof Britain are among the affected
clotting factors which may be inherited or
i ndi vi dual s.
ac quir ed.H e mo p h i l i a ,Vo n W i l l e b ra n d ' sd i sease
etc., are examples of inherited disorder while Hemophilia B (Christmasdisease): This is
afibrinogenemia is an acquired disease. due to the deficiency of Christmas factor (lX).
The cl i ni cal symptomsare al mostsi mi l arto that
Hemophilia A (classicalhemophilia) : This is
found i n hemophi l i aA .
a sex-linked disorder transmitted bv females
affecting males. Hemophilia A is the most Von Willebrand's disease: This disorder is
common clotting abnormalityand is due to the characterizedby failure of plateletsto aggregate
deficiency of antihemophilic factor (VIil). The and is due to a defect in the olateletadherence
affectedindividualshave prolongedclottingtime factor.

BIoMEDTCAL/ GUNTCALCONCEPTS

rx' Albumin, the most abundant plasma protein, is inuolued in osmotic t'unction,
transport of several compounds (fqtty actds, steroid hormones), besides the bulfering
action.
!ii Hypoalbuminemia qnd albuminuria are obseruedin nephrotic syndrome.

uyAntitrypsin deliciency has been tmplicated in emphysema (abnormal distension ol

lungs by qir) which is more commonly associoted with heauy smoking.

Haptoglobin preuents the possible loss of free hemoglobin trom the plasma through the
kidneys by lorming hoptoglobin-hemoglobin complex.
[.t" Immunoglobulins (antibodies), a specialized group of plasma globulor proteins, are
actiuely inuolued in immunity. lgG and lgM are primarily concerned with humoral
immunity while IgE is ossociofed with allergic reactions.

Multiple myeloma, a plasma cell cancer diseoseof bone marrow, is characterlzedbg

ouerproduction ol abnormal immunoglobulins (mostly lgG). Laborotory diognosis ol
multtple myelomo can be made by the presenceof a distinct M band on plasmo/serum
electrophoresis,
I i1
Blood clotting or coagulation is the body's major d.efense mechanism against
blood loss. Delects in clotting factors cause coagulation abnormalities such
as hemophllia A (det'lciency of lactor VIII) qnd Christmos diseose (deficiency of
foctor IX).
st Anticosgulants Inhibit blood clotttng. These include heparin, qxalate, fluoride, EDTA
and citrate.
 B IOC H E MIS TR Y
194

.a '1F.ftt1iI";F..q'

6-8 g/dl. Electrophoresisseporotes

L The total concentrationof plosma proteins is about
a1, &2, p and y globulins'
plasma proteins into 5 distinct bqnds, namely albumin,

2 ' A l b u mi n i s th e m a j o rc o n s ti tu ent(60% )oJpl osmoprotei nsw i thaconcentrati onS ' 5to

5 .0 g /d | .Iti s e x c l u s i u e l y s y n thesi zedbgthel i uer,A l bumtnperformsosmoti c,transpor t
ond nutritiue functiorts.
deficiencv
3. ayAntitrgrpsinis a major constituent ot'.al globul^,!liil',"i^^?:-Antitrvpsin
liuer dtsease'
his been implicated in emphysemaand a speciJic

4 ' H a p to g l o b i n (H p )b i n d s w i thfreehemogl obi n(H b)thatspi l l si ntothepl asmadueto

the glomeruli, hence haptoglobin
hemolysis.The Hp-Hb complex connot pass through
preuentsthe loss of t'ree hemoglobininto urtne'
ceruloplasrnin' C'reoctiue
5. Alterations in the acute phase proteins (e.g' ayantitrypsin,
response to the stimulus of infection'
protein) are obseruedos o result oJ non-spicit'ii
i n j u ry ,i n fl o mma ti o n e tc ' Esti mati onofserumC -reacti ueprotei ni susedforthe
euqluotionot' acute phase response'
the body ogoinst the forergn
6. Immunoglobulins ore specialized proteins to defend
subsfonces.They are mostlg associatedwith yglobulin t'ractionof plasmaproteins' The.
two ilentical heauy choins and two identical
immunoglofullinsessenfiollg consist ol
Iight chains, held together by disult'ide linkages'
IgA, lgM., IgD.and lgE-<re t'ound-in
7. Fiuecrossesof immunogroburins-namery IgG,
immunitg. lgA
humans.IgG is most aiundant and is ^ioi"tv reiponsiblet'or .humoral while
lgM seruesos a t'irst tine oi d"1"nt" t'or humoral immunitg
protects bodg surt'aces.
IgE is associated with allergic reactions'

S.Mu l ti p | e mg e l o m a i s d u e to t hemal i gnancyot' asi ngl ecl o-ne^ot' pl asmacel l si nthebon e

lgG- plosma of multiple
marroLu.This couses the ouerproduction oJ abnormal .The
M-band'
myeloma patients on electrophore-sisshous a distinct
againstblood loss' The extrinsic
9. Blood clotting is the bodg's maior defense mechanism
xa which then participotes in .the
and intrinsic pathways lead to the formation of factor
prothrombin to thrornbin' Fibrinogen is then
final common pathway to actiuate
conuertedto librin clot.
of fibrtn clots. Plasminogen,
lO. Plasmin is mosfly responsible Jor the dissolufion
the inactiue precursor of plasmin' Tissueplosminogen
synthesizedby the kidieg, is
plasmin'
ictiuator (TPA) and urokinase conuert plosminogen to