4) Protein Denaturation

• All proteins after being synthesized must fold during and following synthesis to take up its native conformation.
• The loss of protein structure results in loss of function.
• A loss of three-dimensional structure sufficient to cause loss of function is called denaturation.
• The denatured state is not necessarily a state of complete unfolding of the protein.
• Most proteins can be denatured by heat, which affects the weak interactions in a protein (primarily hydrogen bonds).
• The very heat-stable proteins of thermophilic bacteria may function at the temperature of hot springs (~100 C).
• The proteins can be denatured also by extremes of pH, by certain organic solvents (alcohol or acetone), by certain solutes such as urea and guanidine hydrochloride, or by detergents.
• Organic solvents, urea, and detergents act primarily by disrupting the hydrophobic interactions that make up the stable core of globular proteins.
• Extremes of pH alter the net charge on the protein, causing electrostatic changes and disruption of some electrostatic and hydrogen bonding.
• Amino acid sequence (primary structure) determines tertiary structure.
• Nephosis syndrome is characterized by increased total protein, Albumin, a2-Globulin, and g-Globulin.
• Advanced cirrhosis is characterized by increased total protein, Albumin, and g-Globulin.
• a1-Globulins levels in blood range from 1.2-3.0 g/l.
• Plasmacytoma is characterized by normal total protein, increased Albumin, and g-Globulin.
• a2-Globulins levels in blood range from 3.3-7.3 g/l.
• Globulins are antibodies and are part of the immune defense.
• Acute inflammation or exacerbation of chronic inflammation is characterized by normal total proteins, but increased Albumin, a2-Globulin, and g-Globulin.
• g-Globulins levels in blood range from 7.2-15.6 g/l.
• b-Globulins levels in blood range from 5.5-9.5 g/l.
• Albumin levels in blood range from 40.3-57.7 g/l.
• The most important proof of this is the reversible denaturation of some proteins in some conditions- This process is called renaturation.
• b Globulins include Hemopexin, C-reactive protein (CRP), Transferrin, and LDL (b).
• a2 Globulins include Ceruloplasmin, Haptoglobin, a2 Macroglobulin, Protrombin, and VLDL (pre-b).
• Albumin is the most abundant plasma protein, regulates the osmotic and oncotic blood pressure, and is involved in the transportation of hydrophobic organic anions such as free fatty acids (FFA), bilirubin, aldosterone, and water-insoluble compounds (drugs).
• a1 Globulins include thyroxine-binding protein, retinol-binding protein, a1 Antiproteinase (a1 Antitrypsin), and Transcortin (corticosteroid-binding globulin).
• Prealbumine binds and transports thyroxine and retinol-binding protein.
• Cellulose acetate gel electrophoresis of plasma proteins is a method used to separate plasma proteins into five bands, designated albumin, a1, a2, β, and γ fractions, respectively.
• Its use permits resolution, after staining, of plasma proteins into five bands, designated albumin, a1, a2, β, and γ fractions, respectively.
• The folding pathway of a large polypeptide chain is complicated.
• The folding process occurs as a stepwise process: Local secondary structures are formed first, followed by longer-range interactions between two helices that come together to form stable supersecondary structures, and the process continues until complete domains form and the entire polypeptide is folded.
• Some proteins undergo assisted folding- not all proteins fold spontaneously.
• Folding for many proteins is facilitated by the action of specialized proteins, called Molecular chaperones.
• Molecular chaperones are proteins that interact with partially folded or improperly folded polypeptides, facilitating correct folding pathways or providing microenvironments in which folding can occur.
• The pH at which the complex charge of the protein molecule is 0, also known as isoelectric point (pI), is a crucial factor in understanding the behavior of proteins in electrophoresis.
• In SDS-PAGE (SDS polyacrylamide gel electrophoresis), the proteins are separated basically on their molecular mass (molecular weight).
• The polyacrylamide gel acts as a molecular filter, slowing the migration of proteins approximately in proportion to their charge-to-mass ratio.
• At pI, the molecule is electrically neutral and does not move (migrate) in electrophoresis.
• SDS binds to most proteins in amounts roughly proportional to the molecular weight of the protein, about one molecule of SDS for every two amino acid residues.
• Two classes of molecular chaperones have been well studied: a) called Hsp70; and b) chaperonins.
• The separation of individual proteins from a complex mixture is frequently accomplished by the use of electrolytes.