Biochemistry.

Protein and
Lipids
Nino Gulatava
 Proteins
Proteins are found in all the cells and fluid of the body. They are
synthesized within the cells of the body and released into the
intestinal fluid of the tissue, and the into plasma.
Classification:
1. Physicochemical properties – solubilities: globular proteins are
water soluble, fibrous proteins- not water soluble.
2. Conjugated or simple protein. The conjugated proteins consist of
two components- apoprotein and nonprotein prosthetic group.
These specific prosthetic group may be a carbohydrate
(glycoproteins mucoproteins), a lipid ( lipoproteins), a metal ion
(metalloproteins), a phosphates (phosphoproteins). Simple
proteins – Globular and Fibrous.
Proteins
Proteins-synthesis, metabolism, catabolism.
Most proteins with the exception of some immunoglobulins and proteins hormones, are synthesized
in the liver. Albumin, alpha-globulins, beta-globulins and some gamma-globulins are secreted by the
hepatocytes and enter the general circulation via central veins of the liver. Most immunoglobulins
originate from plasma cells in the bone marrow. Synthesis of proteins occurs as a result of covalent
linking of amino acids. The amino acid sequence is predetermined by the genetic code within the
cell. Deoxyribonucleic acid is transcribed into messenger RNA, which becomes translated in the
cytoplasm of the protein-synthesizing cells. Proteins synthesis then occurs at a rate of approximately
2 to 6 peptide bonds per sec. Synthesis of intracellular proteins is not only controlled by genetic
code, but is also influenced by certain anabolic hormones ( thyroxine, insulin, growth hormone,
testosterone and etc.)
Plasma proteins circulate both in the intravascular and extravascular fluid compartments. The
movement occurs by passive diffusion and active transport.
Catabolism of plasma proteins occurs in the liver. Following hydrolysis of the protein, removal of
the amino group from the amino acids occurs within the hepatocytes. Subsequently, ammonia and
keto acids are formed. Ammonia is detoxified by conversation to urea and is excreted in the urine.
The keto acids enter in the Krebs cycle and are recycled to amino acids or become converted to
glucose or fat to be utilized for fuel. Catabolism of proteins is promoted by the presence of glucagon
and cortisol.
In healthy, nondiseased individual, a balance exists between protein anabolism and catabolism.
Nitrogen balance, the difference intake and excretion of nitrogen, is one of the most widely used
indicators of assessing relative protein change. In healthy individuals, anabolic and catabolic rates
are in equilibrium and nitrogen balance is zero.
Proteins
Protein
The main method to measure total protein is Biuret method, when
formation of a violet-colored complex when Cu ions in alkaline
solution bind the peptide bonds of the protein, maximum absorbance
occurs at 540 nm.
Normal value of Total protein is 66 – 87 g/L.
Concentrations of total protein depends of synthesis and destroy of
two main protein fractions- albumin and globulins.
Evaluation of total protein is hyperproteinemia
Decrease of total protein is hypoproteinemia.
Hyperproteinemia
• Polyclonal or monoclonal gammopathies ( paraproteinurias)
• Certain chronic disease
• Chronic inflammatory conditions disease
• autoimmune disease
• Systemic lupus erythematosus
• sarcoidosis
Protein

Hypoproteinemia:
• Synthesis of proteins are in main cells of liver and in reticuloendothelial
systems
• Increase of protein loss : several disease of kidney, bleeding, diabetes mellitus,
ascites, burns
• disorders of proteins synthesis :hepatitis, cirrosis, toxic disorders, prolonged
treatment by corticosteroids, enteritis, enterocolitis, pancreatitis
• prolonged starvation or prolonged unprotein diets
• Hyperproteinemia:
• dehydration – trauma, infection disease, burns
• there are appears of paraproteins - myeloma, Valdestreme disease.
Protein
Albumin
Normal value – 34 – 48 g/L.
Albumin is 60 % of total protein.
Main function of albumin is transport of hormones, cholesterol,
bilirubin, calcium, some of medicines.
Hyperalbuminemia - dehydration – trauma, infection disease, burns.
Hypoalbuminemia – reasons are the same as the hypoproteinemia.
Separation of proteins
One of the simplest techniques for assessing abnormalities of serum proteins is to perform serum
protein electrophoresis (SPE).
Electrophoresis is based on the principle of separation of proteins when subjected to
electromagnetic field. Since proteins are amphoteric and carry either a positive or negative net
charge, they migrate toward the cathode or anode in a electrophoretic system.The rate of migration
depends upon the following factors:
1. Net charge of protein
2. Size and shape of the protein
3. Electric field strength of the system.
Movement of the protein is depend on the degree of ionization of the protein at the pH of the
buffer.
The most popular used buffer systems for electrophoresis are: barbial, TRIS, TRIS-boric acid-EDTA.

Buffers of an alkaline pH (8.6) are used for serum protein electrophoresis. Serum protein separate
into five bands or zones. In order of the fastest to the slowest moving fractions, the bands appear as
albumin, alpha1-globulins, alpha2-globulins, beta-globulins, gamma-globulins.
Serum protein electrophoresis
The procedure of electrophoresis involves placing the support
medium containing the patient sample into an electrophoresis
chamber, which contains the appropriate buffer. Separation occurs
while a constant voltage is applied for a predetermined period of
time. The electrophoric strip is then removed and proteins are
rapidly fixed, stained, and dried.
Electrophoretic patterns yield both qualitative and quantitative
information. Relative increases and decreases of each protein fraction
can be noted, as well as the homogenicity of each band.
Serum protein electrophoresis

Prealbumin – 2- 7 %
• Albumin – 52 – 65 %
• Alfa- 1-globulin – 2.5 – 5% (alfa-1 antitripsin, alfa-1 lipoproteid,
alfa-1 – glycoproteid).
• Alfa – 2 –globulin – 7 – 13 % ( alfa-2 macroglobulin, haptoglobin,
apoliporotein A, B, C, ceruloplasmin).
• Beta – globulin - 8- 14 %(transferrin, hemopepsin, components of
compliments, Ig and lipoproteins).
• Gamma – globulin – 12 – 22%.(Ig G, A, M, D, E).
Serum protein electrophoresis
• Prealbumin – decrease of level is early and sensitive test of protein
deficiency.
• Elevation of Alfa- 1-globulin – acute inflammatory process, liver
damage.
• Elevation of Alfa- 2 -globulin – acute inflammatory process, especially
with exudative and purulent process (pneumonia, empyema of pleura),
collagenases, autoimmune diseases, cancers and etc.
• Elevation of Beta-globulin – hyperlipidemia, liver diseases, nephrotic
syndrome, hypothyreosis.
• Elevation of gamma –globulins – synthesis of antibodies: viral and
bacterial infections, collagenases, inflammatory processes, chronic
hepatitis.
Lipid
Fats (or triglycerides) within the body are ingested as food or synthesized by
adipocytes or hepatocytes from carbohydrate precursors. Lipid metabolism
entails the oxidation of fatty acids to either generate energy or synthesize new
lipids from smaller constituent molecules. Lipid metabolism is associated with
carbohydrate metabolism, as products of glucose (such as acetyl CoA) can be
converted into lipids.
Lipid metabolism begins in the intestine where ingested triglycerides are
broken down into smaller chain fatty acids and subsequently
into monoglyceride molecules by pancreatic lipases, enzymes that break down
fats after they are emulsified by bile salts. When food reaches the small
intestine in the form of chyme, a digestive hormone called cholecystokinin
(CCK) is released by intestinal cells in the intestinal mucosa. CCK stimulates
the release of pancreatic lipase from the pancreas and stimulates the
contraction of the gallbladder to release stored bile salts into the intestine.
CCK also travels to the brain, where it can act as a hunger suppressant.
Lipid
Together, the pancreatic lipases and bile salts break down triglycerides
into free fatty acids. These fatty acids can be transported across the
intestinal membrane. However, once they cross the membrane, they
are recombined to again form triglyceride molecules. Within the
intestinal cells, these triglycerides are packaged along with cholesterol
molecules in phospholipid vesicles called chylomicrons. The
chylomicrons enable fats and cholesterol to move within the aqueous
environment of your lymphatic and circulatory systems. Chylomicrons
leave the enterocytes by exocytosis and enter the lymphatic system via
lacteals in the villi of the intestine. From the lymphatic system, the
chylomicrons are transported to the circulatory system. Once in the
circulation, they can either go to the liver or be stored in fat cells
(adipocytes) that comprise adipose (fat) tissue found throughout the
body.
Lipid classes
1. Sterol (cholesterol)
2. Triglycerides
3. Phospholipids
4. Fatty acids
Function
Cholesterol- cellular membrane structure and a metabolic precursor for other
biological steroids.
Triglycerides- primary constituents for long-term energy storage. 1g TG
oxidized to CO2 and H2O produced 9 kcal od energy as compared to 4 kcal from
a 1g glycogen.
Phospholipids- one of the principal components of cellular membranes.
Fatty acids – minor constituent in circulating plasma lipids or tissue stores.
Lipoprotein terminology
Electrophoretic mobility Ultracentrifuge fractions
Chylomicron Chylomicron
Pre-beta Very Low Density Lipoprotein
Beta Low Density Lipoprotein
Alpha High Density Lipoprotein
Lipid profile

Atherogenic hyperlipoproteinemia will be appears as results of gens

anomalies, which regulate functions of receptions, enzymes and
transport proteins, which are participate in lipid metabolism. In these
cases there are family disorders of lipid metabolism. But usually we
have primary polygenic hyperlipoproteinemia as results combination
of genetic factors with external environment :smoking, malnutrition,
sedentary lifestyle.
Currently we are use Fredrickson’s classification of
hyperlipoproteinemia , which is confirmed in 1970.According this
classification there 6 type of hyperlipoproteinemia. In real practice
we are meet with IIa, IIb and IV types.
Total Cholesterol

Cholesterol we receive from foods, but most of part is synthesized in

liver. Cholesterol is membrane component of cells, predecessor of
steroid hormones and bile acids.
About 10% of population have hypercholesterolemia, but this fact
proceeds latently. Together with Triglycerides Total cholesterol gives
important information about lipid profile, future algorithm for
diagnostic of lipid’s disorder, hospitalization, choice of treatment
method and than monitoring of medicaments.
There are addiction of elevation Total Cholesterol level and risk of
development ischemic heart disease.
If patient has risk of ischemic disease is recommend to measurement
T-CHOL every 3 month.
Total Cholesterol
< 5.0 mmol/L low risk of atherosclerosis

5.0 – 5.9 mmol/L moderate risk of atherosclerosis

>6.0 mmol/L high risk of atherosclerosis

Hypercholesterinemia
• Hyperlipoproteinemia IIa, IIb ,IV, V, and also III,I
• Liver disorders
• Intra- and extra hepatic cholestasis
• Cancer of pancreas and prostate
• Glomerulonephritis
• Hypothyreosis
• Nephritic syndrome
• Alcoholism
• Hypertonic disease
• Ischemic heart disease
• Diabetes mellitus
• Podagra disease
• Analbunemia and disglobuliemia
• Medicaments: corticosteroids. Corticotrophin, adrenalin, sulfanilamids, thyazid diuretics
Hypocholesterinemia
• Hypoproteinemia and abetalipoproteinemia
• Cirrhosis of liver
• Liver cancer
• Hyperthyreosis
• Malabsorption syndrome
• Thalassemia, megaloblastic anemia
• Chronic disease of lung
• Rheumatoid arthritis
• Rapid fall of level T-CHOL in liver disease is bad prognostic
symptom.
Triglicerides

Normal value : < 1.7 mmol/L

This is neutral fats. Triglycerides we receive as from foods and as
synthesized in liver from carbohydrates. Triglycerides is main forms
of accumulation fats acids in our body and reserve source of energy
during of starvation, long physical activity.
1.71 – 2.27 mmol/L moderate increase

2.28 – 5.69 mmol/L high increase

≥5.7mmol/L significant high increase

LDL – Cholesterol
(Low density Lipoprotein or beta Cholesterol)

Normal value: < 3.37 mmol/L.

This lipoprotein is main transport forms of cholesterol.
Currently we are use classification:
< 3.0 mmol/L low risk of atherosclerosis

3.0 – 3.9 mmol/L moderate risk of atherosclerosis

>4.0 mmol/L high risk of atherosclerosis

LDL – Cholesterol
This parameter more closely correlate with risk of development of
atherosclerosis .
LDL is transport 2/3 of plasma cholesterol . Size of this compounds (d 21-
25 nm) allows as HDL permit vessel wall across endothelial barrier, but
unlike HDL , LDL delay in the wall of vessel, because has select affinity
to glycosaminoglycans and smooth muscle cells. Affinity to
glycosaminoglycans and smooth muscle cells explained in one hand
presence in LDL-CHOL Apo B, and other hand – presence of vessel wall
cell’s surface a receptor to apolipoprotein B. According these facts, LDL is
transport protein for vessel walls , but in pathology conditions - source
of accumulate himself in the wall of vessels.
Measurement of this test very important and informative for estimate of
risk developing atherosclerosis. Concernig this, treatment hyperlipidemia
by medicaments based on the level of LDL-CHOL.
VLDL – Cholesterol (Very Low density Lipoprotein or pre-beta Cholesterol)

Normal value: 0.26 – 1.04 mmol/L.

These lipoproteides are synthesed in liver and is an main transport
forms for triglycerides. Increase of level VLDL-Chol always correlate
with increase level of triglycerides.
VLDL= TG/2.18 mmol/L. (this formula is use when TG < 4.5 mmol/l).
A separate measurement of VLDL have not diagnostic value.
HDL – Cholesterol
(High density Lipoprotein or alpha Cholesterol)

Normal value: > 1.45 mmol/L.

HDL is carry out transport cholesterol from peripheral organs to liver
where cholesterol transformed in bile acids and excreted from the
body.
< 1.04 mmol/L high risk of atherosclerosis

≥1.0 mmol/L moderate risk of atherosclerosis

>1.45 mmol/L low risk of atherosclerosis

HDL – Cholesterol
Increase:
• Primary biliary cirrhosis of liver
• Chronic hepatitis
• Alcoholism
• Intoxications
Decrease:
• Diabetes mellitus
• Kidney and liver diseases
• Hyperlipoproteinemia IV types
• Acute bacterial and virus infections
Electrophoretic mobility
One of method to determination of lipoproteins is electrophoresis.
Analysis using this method each fraction of lipoproteins has
electrophoresis transmission and than these fraction of lipoproteins’
compare to transmission other serum proteins. Based on electrophoresis
lipoproteins divided by:
• Alpha-lipoproteins. During electrophoresis they move with alpha
globulins and are consistent to HDL-cholesterol. Contein:50%
proteins, 30 % phospholipids and 20 % cholesterol, small amount
triglycerides.
• Beta-lipoproteins. During electrophoresis they move with beta
globulins and are consistent to LDL-cholesterol. Contain: 50%
cholesterol,20 % phospholipids and 8-10% triglycerides.
• Prebeta-lipoproteins. Consistent to VLDL-cholesterol.
Apolipoproteins
Apo-A-1protein in serum.
Normal value: male – 1.04-2.02 g/L: female – 1.08 – 2.25 g/L.
Each lipoprotein characterized by presence in composition individual
protein – apoprotein.Apo-A-1 is main apolipoprotein of alpha
lipoproteins.
A separate measurement have not diagnostic value.
Apo-B protein in serum.
Normal value: male – 0.66 – 1.336 g/L: female – 0.6 - 1.17 g/L. This
parameter is main apolipoprotein of beta lipoproteins.