Proteins

• What are proteins composed of?

  Amino acids linked via peptide bonds
• What is the role of proteins in the body?

  They provide oncotic pressure and act as transport and storage molecules
• What is the total plasma protein concentration range?

  60 – 70 g/L
• What are the 20 primordial amino acids categorized into?

  • Nonpolar (9): glycine, alanine, tryptophan, valine, leucine, methionine, isoleucine, phenylalanine, proline
  • Uncharged Polar (6): serine, threonine, tyrosine, asparagine, cysteine, glutamine
  • Acidic, negatively charged (2): aspartic acid, glutamic acid
  • Basic, positively charged (3): histidine, lysine, arginine
• What is the isoelectric point (pI) of an amino acid?

  It is the pH at which the amino acid has no net charge and exists as a zwitterion
• What is the albumin concentration in plasma?

  32 – 45 g/L
• What is the globulin concentration in plasma?

  23 – 35 g/L
• What is the fibrinogen level in plasma?

  2 – 4 g/L
• What does the albumin:globulin (A/G) ratio indicate?

  It indicates the relative concentrations of albumin and globulins in plasma
• What is the conversion factor from g/dL to g/L?

  10
• What is the Kjeldahl Digestion Method used for?

  It is the standard reference method for the measurement of proteins
• How does the Berthelot method measure ammonia?

  Ammonia reacts with alkaline hypochlorite to form a blue product
• What is the purpose of the Nesslerization method?

  It detects ammonium ions using Nessler’s reagent to form a complex
• What does the Biuret Reaction involve?

  It involves the formation of purple complexes that can be measured spectrophotometrically
• At what wavelength is the purple complex measured in the Biuret Reaction?

  540-560 nm
• What does the Lowry Protein Assay combine?

  It combines the Biuret reaction followed by the Folin-Ciocalteau method
• What is the Coomasie Brilliant Blue Method used for?

  It is a sensitive and highly selective method of protein measurement in cerebrospinal fluid
• What does the Ninhydrin Method detect?

  It detects peptides as violet products after separation of amino acids
• What does refractometry measure in relation to proteins?

  It checks the influence of proteins on the refractive index of the samples
• What is the purpose of the UV Absorption Method?

  It measures protein concentration based on UV light absorption at 280 nm
• What is the significance of Polyacrylamide Gel Electrophoresis (PAGE-SDS)?

  It separates proteins by their differences in molecular weight and net isoelectric points
• What are the standard dyes used for qualitative characterization of proteins?

  Coomassie brilliant blue, Ponceau S, Amido black, and silver staining
• What does the absence of an albumin band in PAGE-SDS indicate?

  It indicates analbuminemia
• What is the role of prealbumin?

  It is an indicator of nutrition and binds thyroid hormones
• What is the primary function of albumin?

  It binds bilirubin, steroids, and fatty acids, contributing to oncotic pressure
• What is the function of α1-Antitrypsin?


  It is a major α1-globulin and a protease inhibitor
• What does ceruloplasmin contain?

  Copper
• What is the role of transferrin?

  It transports iron (III)
• What does C-reactive protein (CRP) indicate?

  It is an acute-phase reactant that motivates phagocytosis in inflammatory disease
• What are the reference ranges for serum protein electrophoresis constituents?

  • Albumin: 52 – 68%
  • α1-globulin: 2.4 – 5.3%
  • α2-globulin: 6.6 – 13.5%
  • β-globulin: 8.5 – 14.5%
  • γ-globulin: 10.7 – 21.0%
• What are the conditions causing abnormal serum protein electrophoresis?

  • Hypoalbuminemia: Low spike for albumin
  • Chronic liver disease: Large elevation of γ-globulins
  • Chronic inflammatory disease: Small elevation of γ-globulins
  • Dysproteinemia: Elevated spike in γ-globulins
  • Hypogammaglobulinemia: Flat curve for γ-globulins
  • Nephrotic syndrome: Low albumin spike, high α2-globulins
• What are the conditions causing hypoalbuminemia?

  • Reduced synthesis: Malnutrition, chronic liver disease, genetic abnormalities
  • Increased loss: Nephrotic syndrome, massive burns
  • Increased catabolism: Massive burns, widespread malignancy
  • Multifactorial: Cirrhosis, congestive heart failure
• What conditions elicit C-reactive protein?

  • Almost Always Present: Rheumatoid fever, acute bacterial infections
  • Sometimes Present: Multiple sclerosis, postsurgical state
  • Frequently Present: Active tuberculosis, advanced malignant tumors
• Each amino acid has a characteristic isoelectric point (pI). It has a double charge therefore zwitterionic, and can act as a base or an acid thus ampholytes.
• Total protein (TP) is composed of

  albumin and globulins.
• albumin : globulin ratio (A/G ratio)

  1.3 – 3.0 : 1
• Using the Kjeldahl Digestion Method, proteins can be

  digested into nitrogen compounds in the form of

  ammonia by oxidation at


  350C using sulfuric acid
• Berthelot method (Indophenol reaction)

   ammonia reacts with alkaline hypochlorite to form

  a blue product

   Sodium nitroprusside serves as catalyst
• Nesslerization

   ammonium ions reacts with Nessler’s reagent (2

  HgI2 2 KI) in the presence of catalysts such as

  copper sulfate, selenium or mercury to form

  the Nessler’s complex – HgOHNHHgI.

   Gum ghatti stabilizes the reaction
• Titrimetry

   ammonia in solution is titrated with an acid