Kursk state medical university

• Name:Dalhatu Saidu

• Group:31

• Year:2nd year 4th

semester

Department:biochemistry

BLOOD PLASMA PROTIENS

CONTENTS:

• 1.CHEMICAL COMPOSITION OF BLOOD

• 2.PROTIENS OF BLOOD PLASMA(METHOD OF SEPERATION,CLASSIFICATION OF BLOOD PROTIENS).

• 3.ALBUMINS(PROPERTIES AND FUNCTION).

• 4.α-globulins,β-globulins,ϒ-globulins.

• 5.ENZYMES OF BLOOD PLASMA(diagnosis of diseases by means of enzymes).

• 6.LIPOPROTIENS OF BLOOD PLASMA.

• 7.NITROGEN-CONTAINING COMPOUNDS OF BLOOD PLASMA.

• 8.Non-PROTIEN NITROGEN COMPOUNDS OF BLOOD.

• 9.AZOTEMIA(REASONS FOR DEVELOPMENT,TYPES,COMPENSATION).

COMPOSITION OF BLOOD

When formed elements are removed from blood, a straw coloured liquid

called blood plasma is left. The table below describes the chemical

composition of blood plasma-

PLASMA

WATER(91.5%)Liquid portion of blood. Acts as solvent and suspending medium for

components of blood; absorbs, transports and releases heat.

PLASMA PROTEIN(7.0%)

Exert colloid osmotic pressure , which helps maintain water balance between blood and tissues and regulates blood volume.

ALBUMINSmallest and most numerous blood plasma proteins; produces by liver. Transports proteins for several steroid hormones and for fatty

acids.

GLOBULINSProduces by liver and plasma cells, which develop from B lymphocytes. Antibodies help attack viruses and bacteria. Alpha and beta globulins

transport iron, lipids and fat soluble vitamin.

FIBRINOGENProduces by liver. Plays essential role in blood clotting.

OTHER SOLUTES(1.5%)ELECTROLYTES

Inorganic salts. Positively charges ions(cations) include Na+,K+,Ca+,Mg2+;

Negatively charged ions(anions) include Cl-,HPO42-,SO4

2-,HCO3-.

Help maintain osmotic pressure and plays essential roles in function of cells.

NUTRIENTS

Products of digestion pass into blood for distribution to all body cells. Includes amino acids(from proteins), glucose(from

carbohydrates), fatty acids and glycerol(from triglycerides), vitamins and minerals.

GASESOxygen, Carbon dioxide and Nitrogen. More O2 is associated with

hemoglobin inside red blood cells; more CO2 is dissolved in plasma. N2 is present but has no known functions in the body.

REGULATORY SUBSTANCES

Enzymes, produces by body cells, catalyze chemical reactions. Hormones, produced by endocrine glands, regulate metabolism, growth and development. Vitamins are cofactors for enzymatic

reactions.

WASTE PRODUCTSMost are breakdown products of protein metabolism and are

carried by blood to organs of excretion. Include urea, uric acid, creatine, creatinine, bilirubin and ammonia.

Separation

1. Electrophoresis 2. Ultra-centrifuge

A/G=1.5~2.5

Method of separation of blood plasma protein;

• Plasma proteins are separated by using the inherent differences of each protein. Fractionation involves changing the conditions of the pooled plasma (e.g., the temperature or the acidity) so that proteins that are normally dissolved in the plasma fluid become insoluble, forming large clumps, called precipitate. The insoluble protein can be collected by centrifugation. One of the very effective ways for carrying out this process is the addition of alcohol to the plasma pool while simultaneously cooling the pool. This process is sometimes called cold alcohol fractionation or ethanol fractionation. It was described by and bears the eponym of Dr. Edwin J. Cohn. This procedure is carried out in a series of steps so that a single pool of plasma yields several different protein products, such as albumin and immune globulin. Human serum albumin prepared by this process is used in some vaccines, for treating burn victims, and other medical applications.

Plasma proteins

General characteristics of plasma proteins

1. They are synthesized in liver except

immunoglobulin.

2. Almost all plasma proteins are glycoproteins.

3. Many plasma proteins exhibit polymorphism such

as α1-antitrypsin, transferrin, haptoglobin.

4. Each plasma protein has a characteristic half-life in

the circulation.

5. Acute Phase Proteins, APP

Albumin

Albumin (69 kDa), single polypeptide chain having 585

aa with 17 disulfide bonds, is the most abundant

protein (60%) in the blood plasma. (3.5-5.0 g/dl)

Synthesis of albumin:

– Liver produced about 12g albumin per day which represent 25%

of total hepatic protein synthesis and 50% of secreted protein.

half-life: 20 days

– For this reason, measurement of serum albumin concentration

is used to assays liver function test.

Structure

1. Shape: ellipsoid.

2. Charge: pI=4.0.

3. Domain: three major domains.

Human Serum Albumin

. Transport: It can bind and transport many diverse molecules and serve as low-specificity transport protein, which include:

• a. Metal ions: such as calcium and copper.

• b. Free fatty acid: albumin binds to free fatty acid released by adipose tissue and facilitates their transfer to other tissue.

• c. Bilirubin: this protects from the toxic side effects of unconjugated bilirubin.

• d. Bile acid: albumin carries the bile acids that are recycle from the intestine to the liver in the hepatic portal vein.

• e. Hormones: such as thyroid hormones and the steroid hormones.

• Colloid osmotic pressure, is a form of osmotic pressure exerted by proteins in blood

plasma that usually tends to pull water into the circulatory system.

• Because large plasma proteins cannot easily cross through the capillary walls.

• In conditions where plasma proteins are reduced,

• e.g. from being lost in the urine (proteinuria) or from malnutrition,

• there will be a reduction in osmotic pressure, leading to enhanced fluid retention in

tissue spaces (edema

Function

Colloid osmotic pressure

Low albumin, causing edema.

Clinical aspects

1. Albumin binds different drugs and strongly affects the pharmacokinetics of these drugs.

For example, sulfonamides can

cause the release of unconjugated bilirubin from albumin by competitive binding. If given to infants, sulfonamides may lead to

kernicterus.

2. In cases of liver disease or starvation, albumin synthesis decreases.

This lead to edema.

Clinical aspects

3. Hypoalbuminemia

• lowered plasma albumin

• in malnutrition, nephrotic syndrome and cirrhosis of liver.

4. Albuminuria

• albumin is excreted into urine

• in nephrotic syndrome and certain inflammatory conditions of urinary tract.

5. Albumin is therapeutically useful for the treatment of burns and hemorrhage.

Globulins

α1–Antitrypsin/α1–Antiproteinase(α1–AT or AAT)

It (52 kDa) is a glycoprotein with 394 aa.

It is a major constituent of α1 globulin fraction of

plasma protein, normal concentration about 200mg/dl.

It is a serine protease inhibitor and can combines with

trypsin, elastase and other protease and inhibits them.

Clinical significance1. Emphysema: used to represent the abnormal

distension of lungs.

• About 5% is due to the deficiency of α1–AT.

• This is associated with lung infection and increase

the activity of macrophage to release elastase

that damage lungs tissue.

To methionine sulphoxide and activate alpha att. Smoking can cause oxidation of Met358

2. α1 –antitrypsin deficiency liver disease

due to mutant α1 –antitrypsin accumulates and aggregates to form polymers, by unknown mechanism, cause liver damagefollowed by accumulation of collagen resulting in fibrosis (cirrhosis).

α2 –Macroglobulin (α2 –MG)

It (720 kDa) is a glycoprotein with 4 identical subunits, a major constituent

of α2 fraction.

It is a panprotease inhibitor and can combine and inhibit many protease.

It can bind cytokines such as PDGF and TGFβ and target them to particular cells to

affect on cell growth or function.

Clinical significance

• 2 -MG levels are increased in nephrotic syndrome• a condition wherein the kidneys start to leak out some

of the smaller blood proteins. Because of its size, 2 -MG is retained in the bloodstream.

• This increase has little adverse effect on the health,

but is used as a diagnostic clue.

nephrotic syndromenormal

β or Hepatoglobin (Hp)

It (90 kDa) is a glycoprotein.

It can bind with the free hemoglobin (extra-

corpuscular Hb) in a tight noncovalent complex Hp-Hbduring hemolysis.

Hp-Hb(155 kDa) cannot pass through glomeruli of

kidney while free Hb(65kDa) can and Hp prevent the loss of free Hb into urine.

※Low levels of plasma concentration of Hp can diagnose

hemolytic anemia.

t1/2 of Hp: 5 day, Hp-Hb: 90 min.

ϒ or Immunoglobulins

※Immunoglobulin(Ig)/anti

body(Ab):

※Glycoprotein molecules that

are produced by plasma cells in

response to an immunogen and

which function as antibodies,

mostly associated with γ

fraction.

※But γ-globulin and Ig are not

synonymous.

※Ig is a functional term

※γ-globulin is physical term.

Am

ou

nt

of

pro

tein

Mobility

albumin

globulins

+-

General Functions of Immunoglobulin

1. Antigen(Ag) binding

- Ig binds to a specific antigenic determinant

2. Effector functions

- Complement activation

- Binding to various cells such as phagocytic cells, lymphocytes, mast cells: antibody-mediated phagocytosis or antibody-dependent cell-mediated cytotoxicity (ADCC).

Two Forms of Ig

mIg

1. Membrane Ig, mIg

It confers antigenic specificity on B

cells.

2. Secreted Ig, SIg

It can circulate in the blood and

serve as the effectors of humoral

immunity by searching out and

neutralizing antigens or marking them

for elimination.

SIg

Basic Structure

1. four chains (H2L2): Y shape

two identical light chains (L):

23 kDa

two identical heavy chains (H):

53-75 kDa

2. Disulfide bonds and such

noncovalent interactions as

salt linkages, hydrogen bonds

and hydrophobic bonds to

form heterodimer (H-L).

1. Variable region (V):VL&VH

2. Constant region (C):CL&CH

3. Hinge region: flexibility

Hingeregion

Light Chain:– VL (110 aa) + CL (110 aa)

Heavy Chain:– VH (110 aa) + CH1-CH3 (or

CH4) (330-440 aa)

Structural Regions

In terms of the differences in amino acid sequence of constant region of heavy chain, immunglobulin molecules are divided into 5 classes:

– IgG, IgA, IgM, IgD and IgE

Heavy chain:

– 5 types: γ,α,μ,δ and ε.

Light chains

– 2 types: κand λ.

Immunoglobulin Classes and Subclasses

1. IgG - γ heavy chains

2. IgM - µ heavy chains

pentamer

3. IgA - α heavy chains

dimer

4. IgD - δ heavy chains

5. IgE - ε heavy chains

Immunoglobulin Classes of Mammals

dimer pentamermonomer

IgG

It is the most abundant class in serum, constitutes

about 80% of the total serum Ig.

4 subclasses, IgG1, IgG2, IgG3, and IgG4.

All IgG's are monomers. The subclasses differ in the

number of disulfide bonds and length of the hinge region.

IgG1, IgG2, IgG4 IgG3

Functions of IgG

1. Major Ig in extravascular spaces.

2. Placental transfer: IgG is the only class of Ig that

crosses the placenta.

3. Complement activation.

4. Binding to cells - Macrophages, monocytes, PMNs (polymorphonuclear

leukocyte), and some lymphocytes have Fc receptors for the Fc region of

IgG.

IgA• Structure

• Serum - monomer

• Secretions (sIgA)

• Dimer

• J chain

• Secretory component

J ChainSecretory Piece

IgA• Function• 2nd highest serum Ig

• Major secretory Ig (Mucosal or Local Immunity)• Found in the body secretions: tears, breast milk, saliva, mucus of the bronchial,

genitourinary, and digestive tract

• IgA is the most predominant antibody in the colostrum, the initial secretion from the mother’ breast after a baby is born.

• Does not activate complement (unless aggregated)

• Binds to Fc receptors on some cells

IgM

Structure

The largest Igcomposed of 5 Y-shaped units held together by a J polypeptide chain.

1. Pentamer

2. Extra domain (CH4)

3. J chain

Functions of IgM

1. 3rd highest serum Ig.

2. IgM cannot traverse blood vessels, hence it is restricted to the blood

stream.

3. 1st Ig produced in a primary response to an antigen and serve as first line

of defense.

4. a good complement activation Ig. Thus, IgM is the most effective in

leading to the lysis of microorganisms.

5. Binds to Fc receptors.

IgD

• Structure

• Monomer

• Tail piece

• Properties

• 4th highest serum Ig, its role in serum uncertain.

• B cell surface Ig.

• Does not bind complement

Tail Piece

IgE

• Structure

• Monomer

• Extra domain (CH4)

• Function• Least common serum Ig

• Allergic reactions

Binds to basophils and mast

cells (Does not require Ag

binding)

• Parasitic infections

(Helminthes)• Binds to Fc receptor on

eosinophils

• no complement activation

CH4

Plasma enzymes

Functional Non functional

**For example:-

3-Alanine transminase(ALT) 4-Aspartate transminase(AST)

5-Alkaline phsphatase 6-Acid phosphatase

7-Amylase & Lipase enzymes

Izoenzymes1-Lactate dehydrogenase(LDH)2-Creatine kinase(CK)

Lactate dehydrogenase(LDH)

Lactic acid Pyruvic acid

NAD NADH+H

*LDH is a tetramer(consists of 4 protomers)

*the promoter's are 2 types:-

1-H(after heart)

2-M(after muscle)

so LDH have 5 isoenzymes:-

LDH 1 – Found in heart and red-blood cells and is 17% – 27% of the normal

serum total*It is formed of HHHH.It increases in myocardial infarction

LDH 2 – Found in heart and red-blood cells and is 27% – 37% of the normal

serum total.

*It is formed of HHHM.It increases in myocardial infarction

LDH 3 – Found in a variety of organs and is 18% – 25% of the normal serum

total.

*It is formed of HHMM.It increases in leukaemia

LDH 4 – Found in a variety of organs and is 3% – 8% of the normal serum total.

*It is formed of HMMM.It increases in viral hepatitis

LDH 5 – Found in liver and skeletal muscle and is 0% – 5% of the normal serum

total*It is formed of MMMM.It increases in viral hepatitis

Creatine kinase (CK)

Creatine Ceatine phosphate

ATP ADP

*CK is a dimmer (consists of 2 protomers)

*the protomers are 2 types:

1-B (after brain)

2-M (after muscle)

*so CK has 3 isoenzymes:-

.CPK1 (CPK-BB) is the characteristic isoenzyme in brain and is in significant amounts in smooth muscle and is 0% of the normal serum total.

**It increases in brain tumors.

.CPK2 (CPK-MB) accounts for about 35% of the CPK activity in cardiac muscle , but less than 5% in skeletal muscle and is 0% of the normal serum total.

**It increases in heart diseases.

.CPK3 (CPK-MM) is the predominant isoenzyme in muscle and is 100% of the normal serum total.

**It increases in skeletal muscle diseases.

Alanine transminase(ALT)

*It is also called serum glutamic pyruvic transminase(SGPT)

*ALT is particularly diagnostic of liver involvement as this enzyme is found predominantly in hepatocytes.

*It increases in liver & heart diseases.

*It catalyzes transfer of amino group (NH2) from amino acid (alanine) to a-keto acid producing a new amino acid & a new keto acid

Glutamic oxalacetic transminase(SGOT)

Aspartate transminase(AST)

*It is also called aspartate glutamic oxalacetic transminase(SGOT)

*It increases in liver & heart diseases

*When assaying for both ALT and AST the ratio of the level of these two enzymes can also be diagnostic. Normally in liver disease or damage that is not of viral origin the ratio of ALT/AST is less than1.with viral hepatitis the ALT/AST ratio will be greater than1.

*The level of AST elevation in the serum is directly proportional to the number of cells involved as well as on the time following injury that the AST assay was performed. Following injury, levels of AST rise within 8 hours and peak 24–36 hours later. Within 3–7 days the level of AST should return to pre-injury levels unless further injury occurs.

* Although measurement of AST is not, in and of itself, diagnostic for myocardial infarction, taken together with LDH and CK measurements the level of AST is useful for timing of the infarct

lipoproteins of blood

• Molecular complexes that consist of lipids and proteins. They

function as transport vehicles for lipids in blood plasma.

• Lipoproteins deliver the lipid components (cholesterol and

triglyceride etc.) to various tissues for utilization.

Classification of plasma lipoproteins

according to their density

• Chylomicron (CM)

• Very low density lipoprotein (VLDL)

• Intermediate density lipoprotein (IDL)

• Low density lipoprotein (LDL)

• High density lipoprotein (HDL)

• Each contains different

kinds and amounts of

lipids and proteins

• The more protein,

the higher the

density

• The more lipid, the

lower the density

• Each has different

function

Lipoproteins

Classification of plasma lipoproteins according to

their electrophoretic mobility

(CM)

a-lipoprotein (HDL)

Pre-b-lipoprotein (VLDL)

b-lipoprotein (LDL)

CM

1967, Fredrickson et al.

What do lipoproteins do?

• Serve to transport lipids and lipid-soluble compounds

between tissues and organs

• Substrates for energy metabolism (TG)

• Essential components for cells (PL, C)

• Precursors for hormones (C)

• Lipid soluble vitamins

• Precursors for bile acids (C)

VLDL

• VLDL are made by liver. Liver synthesizes TG and cholesterol and packages them into VLDL for export into blood.

• Most lipid in the core of VLDL is triglyceride

• Nascent VLDL contain apoB100. In blood nascent VLDL pick up apoE and apoCs from HDL and become matured VLDL.

VLDL

• In the capillaries of various tissues, LPL degrades TG to fatty acids and glycerol, which enter the tissues by diffusion. ApoC-II is needed in this step to activate LPL.

• When VLDL loses triglyceride, it transforms into VLDL remnant, also named as IDL (intermediate-density lipoprotein).

• During the process, some apolipoproteins (apo As and apoCs) are transferred back to HDL.

• VLDL function: Deliver TG from liver to peripheral tissue cells.

Fates of VLDL remnants (IDL)

• Results from loss of TG in VLDL

• Contains relatively more cholesterol esters

• Taken up by liver or transform into LDL

1) A proportion of the VLDL remnant (IDL) is

taken up by liver through the LDL receptor

(apoE-mediated).

2) The other remnant is further acted upon by

hepatic lipase (HL) and converted into LDL.

LDL loses all apolipoproteins except apoB100.

VLDL remnant

LDL

Most core lipid in LDL is cholesterol ester.

ApoB100 is only apolipoprotein in the surface.

Non-Protein Nitrogen(NPN)

Compounds

Non-protein Nitrogen Compounds

• The determination of nonprotein nitrogenous substances in the blood has traditionally been used to monitor renal function.

• Nitrogen containing compounds that are not proteins or polypeptides

• Useful clinical information is obtained from individual components of NPN fraction

Clinically Significant NPN• The NPN fraction comprises about 15 compounds

• Majority of these compounds arise from catabolism of proteins and nucleic acids

Clinical Application

•Measurement of urea is used to: • evaluate renal function,

• to assess hydration status,

• to determine nitrogen balance,

• to aid in the diagnosis of renal disease,

• and to verify adequacy of dialysis.

Azotemia

Azotemia characterized by abnormally high levels

of nitrogen-containing

compounds, such as urea, creatinine, various body

waste compounds, and other nitrogen-rich compounds in the blood.

It is largely related to insufficient filtering of blood by the kidneys

Types

1.Prerenal Azotemia

2.Primary Renal Azotemia

3.Postrenal Azotemia

All forms of azotemia are characterized by

a decrease in the GFR of the kidneys

and increases in BUNand serum creatinine

concentrations.

The BUN-to-creatinine ratio is a useful measure

in determining the type of

azotemia.

A normal BUN:Cr

is equal to 15.

GFR

BUN &

Creatinine

15

1. Prerenal azotemia

Prerenal azotemia is caused by a decrease in blood flow

(hypoperfusion) to the kidneys.

It can occur following hemorrhage, shock, volume depletion,

congestive heart failure, and narrowing of the renal artery

among other things.

The BUN:Cr in prerenal azotemia is greater than

20.

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2. Renal azotemia

Renal azotemia (acute renal failure) typically leads to uremia. It is an intrinsic disease of the kidney, generally the result of renal parenchymal damage.

Causes: renal failure, glomerulonephritis, acute tubular necrosis, or any other kind of renal disease.

The BUN:Cr in renal azotemia is less than 15.

In cases of renal disease, glomerular filtration rate

decreases, so nothing gets filtered as well as it normally would.

3. Postrenal azotemia

Blockage of urine flow in an area below the kidneys results in postrenal azotemia.

Causes: vesicoureteral reflux, blockage of the ureters by kidney stones, pregnancy, compression of the ureters by cancer, prostatic hyperplasia, or blockage of the urethra by kidney or bladder stones.

The BUN:Cr in postrenal azotemia is >15.

Signs and symptoms (prerenal azotemia) Oliguria or anuria

Fatigue

Asterixis (flapping tremor)

Decreased alertness

Confusion

Pale skin

Tachycardia

Xerostomia

Thirst

Edema

Orthostatic blood pressure

Uremic frost, a condition that occurs when urea and urea derivatives are secreted through the skin in sweat, which evaporates away to leave solid uric compounds, resembling a frost.

Treatment

Dialysis

Medications to increase cardiac output and increase blood pressure, and the treatment of the condition that caused the azotemia.