PROTEINS

J.L.B.NARVASA, RMT

PROTEINS
These are macromolecules composed of polymers of covalently linked
amino acids that are involved in every cellular processes.
Amphipathic and amphoteric
Composed of 50-70% of the cells dry weight

FUNCTIONS:
Repair body tissues
Important in blood coagulation and immunologic function
For transport of metabolic substances
Maintenance of osmotic pressure
Maintenance of blood pH
Biocatalysts

FOUR PROTEIN STRUCTURE

Primary structure- it is the linear sequence of the amino acid
Secondary structure-it involves the winding of the polypeptide chains
Tertiary structure- the actual 3-dimensional configuration; the folding
pattern of the protein
Quaternary structure- the association of 2 or more polypeptide chains
to form a functional protein molecule.

CLASSIFICATION OF PROTEINS
Simple proteins- are those which are made of amino acid units
only, joined by peptide bond.
Conjugated proteins- are composed of simple proteins combined
with a non-proteinous substance. The non-proteinous substance is
calledprosthetic grouporcofactor.

PLASMA PROTEINS
Prealbumin Haptoglobin
Albumin Hemopexin
Globulin Ceruplasmin
1- Antitrypsin (AAT)

2- Macroglobulin(AMG)

1- Fetoprotein (AFP) 2-Microglobulin

1- Acid Glycoprotein/ orosomucoid (AAG) C-reactive Protein (CRP)
1- Antichymotrypsin Lipoprotein
Group-specific component (Gc)- Globulin
Transferrin
Immunoglobulins
Fibrinogen
Complement

PLASMA PROTEINS
Prealbumin
It migrates ahead of albumin
It is rich in tryptophan and has considerable -pleated sheet conformation
It serves as a transport protein for T4 and retinol
Indicator of nutritional status
It is used as a landmark to confirm that the specimen is really CSF

PLASMA PROTEINS
Albumin
It is the protein present in highest concentration in serum
It is a general transport protein
Indicator of nutritional status
It maintains osmotic pressure
It is a negative acute phase reactant
Lowest plasma levels are seen in active nephrotic syndrome
Reference values= 3.5-5.0 g/dL

PLASMA PROTEINS
Globulin
The globulins are a family of globular proteins that have higher molecular
weights than albumins and are insoluble in pure water but soluble in dilute
salt solutions
Transport protein for steroid and lipid hormones
Consist of four fractions: 1, 2, and fractions
Measurement: Total Protein Albumin = Globulin
Reference values: 2.3-3.5 g/dL

TOTAL PROTEIN DETERMINATION

Kjeldahl Method
Biuret Method
Folin Ciocalteu (Lowry) Method
Ultraviolet Absorption Method
Electrophoresis
Refractometry
Turbidimetric and Nephelometric Methods
Salt Fractionation

TOTAL PROTEIN DETERMINATION

Kjeldahl Method
Standard reference method
It is based on measurement of nitrogen content
It uses serum treated with tungstic acid, forming protein-free filtrate
1 gram of nitrogen- 6.54 grams of proteins

TOTAL PROTEIN DETERMINATION

Biuret Method
Most widely used method
Principle: cupric ions complex the groups involved in the peptide bond
forming a violet-colored chelate which is proportional to the number of
peptide bonds present.
Reagents:
Alkaline copper sulfate
Rochelle Salt (NaK Tartrate)
NaOH and Potassium Iodide

TOTAL PROTEIN DETERMINATION

Electrophoresis
Principle: Migration of charged particles in an electric field
Normal SPE Pattern:
1. Albumin- fastest band
2. Alpha 1- Globulin- 2nd fastest band
3. Alpha 2- Globulin- 3rd fastest band
4. Beta- Globulin- 4th fastest band
5. Gamma- Globulin- slowest band

OTHER MEASUREMENTS FOR PROTEIN

Coomassie Brilliant Blue Dye- for detection down to 1ug of protein
Ninhydrin Test- develops a violet color by reacting with primary
amines
Dyes used for measurement of Albumin
Bromcresol Green(BCG)
Methyl Orange (MO)
Hydroxyazobenzene benzoic acid (HABA)
Bromcresol purple (BCP)

ALBUMIN/ GLOBULIN RATIO

Has been used as an index of disease state, however, it is not a
specific marker for disease because it does not indicate which
specific proteins are altered.
If globulin is greater that albumin, it is known as inverted A/G
ratio.
Reference Value: 1.3-3.1

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