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PLASMA PROTEINS

,
IMMUNOGLOBULINS AND
BLOOD COAGULATION
By
Dr.Hj. Sadiah Achmad,dr

PLASMA PROTEINS, IMMUNOGLOBULINS AND
BLOOD COAGULATION

Blood : - Solid elements : - Red & white blood cells
- Platelets
- Liquid Medium : - Plasma (H2O, electrolytes,
metabolites, nutrients, proteins,
hormones)
Blood has clotted  liquid phase : serum
- Lacks of clotting factors
- Some degradation products of clotting factors

Functions of Blood :
1. Transport :

- O2 & CO2

Respiration

- Nutrients

 Nutrition

- Metabolic waste

Excretion

- Hormones
- Metabolites
2. Homeostasis : - Maintenance of normal acid-base balance
- Regulation of water balance
- Regulation of body temperature
3. Defense : - Immune System - cellular : white blood cells
- humoral : circulating antibodies

PLASMA PROTEINS
 Plasma contains a complex mixture of proteins
- single proteins
- conjugated proteins
 Concentration of total plasma protein : 7.0 - 7.5 g/dL
 Plasma proteins can be separated :
- salting out methods

- Albumin, Globulin, Fibrinogen

- electrophoresis (Cellulose acetate)

bands :

- albumin
- 1, 2, ,  - globulins
Densitometer scanning
bands

graph.

quantified the amount of

 .globulin. 1 -globulin.  .a + – Albumin 1 2   b Albumin 1 2   a : Separated protein bands are visualized in characteristic positions after being stained b : Densitometer scanning from cellulose acetate strip converts bands to characteristic peaks of albumin. 2 .globulin.globulin. .

chronic diarrhea .severe protein malnutrition . . Concentration of plasma protein determine the distribution of fluid between blood & tissues.chronic glomerulo nephritis.Concentration diminished edema due to : • Protein deficiency : . .

glycosylation. phosphorylation) Transit time : 30’ .several hours. Glycoproteins (except albumin) N/O . Study of plasma proteins : a. . secretory vesicles Ser. c. Synthesized on membrane – bound polyribosomes as preprotein Rer. Golgi APP plasma : post translational modifications (proteolysis.g.oligosaccharide chain various functions e. Synthesized in the liver (-globulin in plasma cells) b. removal of sialic acid (from ceruloplasmin) shorten half life.

d.albumin : 20 days .ceruloplasmin.haptoglobin : 5 days Crohn’s disease (regional ileitis) : plasma proteins  lost into bowl through inflamed intestinal mucosa : protein .anti trypsin. Characteristic half life in circulation . Polymorphism.losing gastroenteropathy  half life albumin : 1 day . transferin . . immunoglobulins Strach gel electrophoresis  characteristic migration e.1.ABO blood group substances . haptoglobin.

Haptoglobin.C .Reactive Protein (CRP) : 50% .1. Levels of certain plasma proteins increase : 1. Cancer 1000x . Chronic Inflammatory states 3.anti trypsin. 1.f. Acute inflammatory states / secondary to tissue damage acute phase proteins (reactants) .acidglycoprotein . fibrinogen 2.

Interleukin 1 (1L – 1) : stimulate synthesis of acute phase reactants by hepatocytes (gene transcription) .Response to Inflammation : .CRP : stimulate classical complement pathway .1 – antitrypsin : neutralize proteases released during acute inflammatory states .

Antithrombin . complement proteins .CRP .leakage from cells/tissue : aminotransferases  Immune defense : .Antichymotrypsine.ß2 . Fibrinogen  Enzymes : .1.Coagulation factors.acidglycoprotein (orosomucoid) .cholinesterase.function in blood : .Immunoglobulins.coagulation factors .antitrypsin.  Inflammatory responses : .Functions of Plasma Proteins  Antiproteases : . .microglobulin. 2 .macroglobulin  Blood clotting : .1.

HDL) .Albumin ( various ligands) .Thyroid – binding globulin (T4. estradiol) . LDL.Transferrin (Fe) .Lipoproteins ( CM.Ceruloplasmin (Cu2+) .Retinol – binding protein (retinol) . VLDL.Cortico steroid – binding globulin / transcortin (cortisol) . Transport / binding proteins : .Haptoglobin (extracorpuscular Hb) .Sex hormone – binding globulin (testosteron. T3) .

ALBUMIN  The major plasma protein : 3.7 g/dL 60% of total plasma protein .4 – 4.protein malnutrition (kwashiorkor)  Consist of : 1 polypeptide chain of 585 amino acids & 17 disulfide bonds (69kDa)  Proteases 3 domains : different functions .40% in the plasma .60% in extra cellular space  Liver produces ± 12 g albumin per day 25% of total hepatic protein synthesis  Synthesis is depressed in : . ratio .liver disease decreased albumin / globulin.

Major determinant of plasma osmotic pressure Analbuminemia : mutation that affect splicing moderate edema ( other plasma proteins increase compensate) . bilirubin. Cu2+.Binds various ligands : FFA.burns treatment of . Function : . drugs (sulfonamides.hemorrhagic shock . penicillin G. dicumarol. Ca2+. aspirin)  Preparations of human albumin . steroid hormones.

kidney : Hb is catabolized by liver cells iron is conserved & reused.HAPTOGLOBIN (Hp)  Glycoprotein : 90 kDa  Binds extracorpuscular Hb (Hb .10% of Hb degraded each day circulation : extracorpuscular .90% degraded in RES. Hb kidney in tubules Hb .Hp excreted in urine / precipitates iron is lost.Hp) prevents loss of free Hb into kidney and urine . .

Hp2-2 (no significant functional differences) Starch gel electrophoresis Hp1-1 : single band Hp2-1 Hp2-2 complex band patterns 2 genes direct the 3 phenotypes : Hp1 & Hp2 Hp2-1 : heterozygous phenotype. . Hp2-1. Human Hp : 3 polymorphic forms : Hp1-1.

plasma level .Low level : Hemolytic anemias half life Hp : 5 days (Hb – Hp: 90’) Hb .Hp is an acute phase protein in inflammatory states. The level of Hp : vary diagnostic use .Hp rapidly removed from plasma by hepatocytes cleared from plasma : 80 x faster than normal Hp level falls rapidly in hemolytic anemias .

globulin (glycoprotein). 76 kDa.TRANSFERRIN  ß1. . synthesized in liver  About 20 polymorphic forms  Binds & transports Fe (2 mol Fe3+ per mol of transferrin) internalized by receptor – mediated endocytosis apotransferrin is not degraded within the lysosome reenters the plasma.

Inadequate utilization of Fe . Concentration in plasma : 300 mg/dL bind 300 g/dL Fe total iron binding capacity of plasma Normal : 1/3 saturated with iron Iron def.Excessive loss of Fe . older people ) . . Anemia : less saturated ( < 1/3) Hemochromatosis : > 1/3 saturated Fe deficiency anemia : occurs frequently.Inadequate intake of Fe (pregnancy.

surround 3000 .5 kDa) .FERRITIN * Stores Fe Intracellular * Contains 23% Fe Apoferritin : .4500 Fe 3+ in micellar form * Normal : a little ferritin in plasma In excess Fe : ferritin in plasma Hemochromatosis : ferritin in tissue (liver.440 kDa (24 subunits of 18. spleen) Ferritin in plasma can be measured by Ria (sensitive & specific) index of body iron stores .

* Synthesis of ferritin & transferrin receptor (TfR) : reciprocally linked to cellular iron content.synthesis TfR . Iron Response Elements (specific untranslated sequences of mRNAs for both proteins) interact with IRE. • Fe levels .tfR mRNA : degraded • Fe level .ferritin mRNA inactive .binding protein .ferritin mRNA synthesize ferritin .

HEMOSIDERIN * A partly degraded form of ferritin.repeated transfusions .Excessive oral intake of Fe. . still containing Fe Excessive storage of Fe detected histologically by prussian blue staining ( for Fe) HEMOCHROMATOSIS * Primary CHR : Genetic disorder Excessive storage of Fe in tissue damage * Secondary CHR : .

determination of TIBC & % transferrin saturation . .prussian blue stain of tissue sections .determination of amount of Fe (g/g) in tissue biopsy.determination of ferritin in plasma (RIA) .Lab tests for assessing disorder of Fe metabolism : .RBC count & estimation of Hb .

globulin * Carries 90% Cu in plasma : 1 mol binds tightly 6 atoms Cu (albumin carries 10 % plasma Cu & binds less tightly donates Cu to tissues) * Ceruloplasmin . abnormal metabolism of Cu) .wilson disease (hepatolenticular degeneration).liver disease .CERULOPLASMIN ( 160 kDa) * 2 .

cytochrome oxidase MENKES DISEASE * Disorder of Cu metabolism Mutations in the gene for Cu .amine oxidase. vasculature (liver not involved) * Activation of Cu – dependent enzymes .Cu dependent superoxide dismutase .COPPER (CU) * Essential trace element * Co-factor various enzymes : . connective tissue. tyrosinase .binding P-type ATPase (directing the efflux of Cu from cells) * Accumulation of Cu in nervous tissue.

penicillamine : chelates Cu in urine.liver Cu > 250 g/g .diet low Cu. Neurologic syndrome Kayser-Fleischer ring (green/golden pigment ring around the cornea) Diagnostic : . cirrhosis. Cu fails to be excreted in bile accumulates in liver.plasma ceroluplasmin < 20 mg/dL Treatment : . : copper toxicosis .Hemolytic anemia. . Hepatitis. kidney. RBC. brain. excreted .WILSON DISEASE * Mutations in gene coding Cu – binding P-type ATPase.

 1-ANTIPROTEINASE ( 1-ANTITRYPSIN) * Single chain (52 kDa) : 394 amino acids + 3 oligo saccharide chains * Major component (> 90%) of 1 fraction of plasma * Synthesized by hepatocytes & macrophage * Serin protease inhibitor : inhibiting elastase of neutrophils * 75 polymorphic forms ( can be separated by electrophoresis) * Deficiency 1 . emphysema (5%) .T.A.

A.A.active elastase + proteolysis Smoking : oxidizes met no tissue damage or no 1 .methionine is involved) no proteolysis of lung .T active El.T inactive El : 1 .A. 1 . : .Def.A.T complex (residu 358.T WBC PMN in the lung (pneunomia) lacks acounter- check to proteolytic damage by protease (elastase) : .A. sulfoxide inactive emphysema Treatment of patients with emphysema due to  . tissue damage met.T Def.active elastase + 1 .

A. 1 .hepatocytes hepatitis cirrhosis. .T accumulate & aggregate in the cisternae of end.T : Liver disease Polymers of mutant 1 .A.retic.* Def.

monocytes. . TGF-ß) toward particular tissues / cells. 2 – MACROGLOBULIN * Large plasma glycoprotein : 720 kDa ( 4 identical subunits of 180 kDa) * 8 . hepatocytes & astrocytes * Thiol ester plasma protein family (contain internal cyclic thiol ester bond) * .Targets certain cytokines (PDGF.10% of total plasma protein * Transport 10% of Zn (the rest : by albumin) * Synthesized by .Binds & neutralizes many proteinase panproteinase inhibitor .

cell . T – lymphocytes (thymic) immunologic processes.IMMUNOGLOBULINS (Ig) *  .globulin . glycoprotein * Synthesis : plasma cells (B lymphocytes) * Function : key role in defense mechanism (humoral anti - bodies) Two components of immune system : 1. B – lymphocytes (bone marrow cells) humoral antibodies (Ig) 2.mediated .

Consist of : .2 identical heavy (H) chains (53 – 75 kDa) Held together as tetramer (L2H2) by disulfide bonds (intra & interchain) Y .2 identical light (L) chains (23 kDa) .shaped. CH3 & VH CH2 : complement –binding site CH3 : attaches to receptor on neutrophils & macrophages Antigen – binding site : hypervariable regions of L & H – chains ( located on VL & VH) .CH1. CH2.* Structure . L-chain : 2 domain : .variable region (VL) H-chain : 4 domain : .constant region (CL) .

V –S V C Lc – S H hain –S –S – cha in – –S H C H F.c 1 –S + –S – SS SS – S – – in S a h – – S H c ain –S – h –S Lc H3N –S + H3N Hinge region L -S - H3N –S – -S –S + Fa b L CH2 C H3 –S – S– –S – S– COO- –S – S– –S – S– COO- Pepsin Papain Cleavage sites H chain H chain .S- H3N -S + – S – Fab Structure of IgG .

IgM.* Digestion of Ig by papain (on hinge region) . IgE . IgD.  &  (differences in CH region)  &  : 4 CH domains 5 Ig class : IgG. . . * L – chains : .2 antigen – binding fragments (Fab) : bind 2 mol antigen (on antigenic determinant = epitope) divalent / bivalent . IgA. Ig always contains : 2 or 2  ( chains are more frequent) * H – chains : . .1 crystallizable fragment (Fc). ( differences in CL region) Mol.

does not fix complement .opsonizes bacteria . milk. intestinal & genital tracts.Function : IgG : .fixes complement easier to phagocytose enhances bacterial killing .neutralizes bacterial toxins & viruses . in secretions (saliva.secretary IgA prevents attachment of bacteria & viruses to mucous membrane . tears) & fluids of respiratory. .found in serum .main antibody in the secondary response .crosses the placenta IgA : .

uncertain .mediates immediate hypersensitivity (releasing mediators from mast .IgM : . .does not fix complement.does not cross placenta .antigen receptor on the surface of B cells IgD : .produced in the primary response to an antigen .found in serum & on the surface of many B-cells IgE : .cells & basophils upon exposure to allergen) .defends against helminthic infections (releasing enzymes from eosinophils) .fixes complement .

Light chain hypervariable regions VL Interchain disulfide bonds VH CL CH Intrachain disulfide bonds Heavy chain hypervariable regions Schematic model of an IgG molecule showing approximate positions of the hypervariable regions in heavy and light chains .

L – chains : 3 hypervar.comprise of invariable regions & hypervariable regions . region . r : antigen – binding site & dictate specificity of antibodies complementarity .determining regions (CDRs) : 5-10 amino acids in each CDR .CDRs : located on small loops of V domains surrounding peptide regions between CDRs : frame work regions CDRs from VH & VL : form a single hypervar. surface : antigen – binding site .* Variable Regions of Ig mol .hypervar. heterogenous . region H – chains : 4 hypervar.consist of VL & VH.

transplacental passage .large antigens : interact with all of CDRs . . . e.small ligands : interact with only 1 or a few CDRs that form a pocket or groove in the molecule * Constant regions C.* Various combinations of H & L chains CDRs (combinatorial diversity) different specificities of antibody diversity of antibody molecules.complement fixation .region : CH2.g. CH3 (CH4 of IgM & IgE) : Fc fragment responsible for class-specific effector functions of different Ig mol.

Secretory IgA (dimer) Schematic representation of serum IgA. chain H L Secretory component B.Monomer L H Dimer L H A. secretory IgA .chain L H L L H J. Serum IgA H J.

chain H L H L C.J . IgM (pentamer) Schematic representation of IgM .

Joining region (J) gene .chain : product of 3 structural genes : .VH gene .CL gene H .V L gene .J gene .* IgG : basic tetrameric structure IgA : polymers of 2 –3 tetrameric units IgM : 5 tetrameric units * Both L & H chains : products of multiple genes L .chain : product of 4 different genes : .CH gene .Diversity region (D) gene .

Increased production of specific classes of Ig / specific Ig mol Multiple myloma : tumor of plasma cells Electrophoresis of serum /urine increased of 1 particular Ig / 1 particular L chain (Bence Jones Protein) .g.Decreased production : • single class of Ig mol ( e. IgA or IgG) a gamma globulinemia : genetic abnormality of IgG impairment of the body’s defense against microorganism (immunodeficiency) • all classes of Ig .* Disorders of Ig .

other process * .generation of peptide fragments various aspects of inflammation : chemotaxis. .comprises 20 plasma proteins . inflammation.The Complement System * Involved in : .cell lysis.major protein components : C1-9.cell lysis . C9 associated with C5 – 8 complex membrane attack complex : generating a lipid-soluble pore in cell membrane osmotic lysis * Detail of the system : complex Basic concept : Inactive protein of the system stimulated activated (by proteolysis) interact with other proteins of the system result : . phagocytosis etc.labile when heated at 56oC .

clearance of antigen .Other function of the system : .antibody complexes from circulation * Activation of the system : .alternative pathway : direct interaction of bacterial cell surfaces (polysaccharides) with C3b ( not involving antibody) .classic pathway : interaction of C1 with antigen – antibody complexes .