Proteins and Denaturation: Plasma Proteins

• This covers protein denaturation, purification, and plasma proteins.

Protein Denaturation

• Protein denaturation involves changes to a protein's structure.
• This includes its definition, causes, effects, and clinical relevance.

Definition

• Protein denaturation is the loss of a protein's functional structure, except for its primary structure.

Causes

• Factors that lead to protein denaturation include heat, pH changes, heavy metals, detergents, mechanical mixing, and digestive enzymes.

Effects

• Chemical effects of denaturation include the loss of most bonds and the loss of peptide bonds through digestive enzymes.
• Physical changes involve decreased solubility and increased viscosity.
• Biologically, denaturation leads to a loss of function, like enzymes ceasing to work.

Clinical Relevance

• An example of denaturation's clinical relevance is the heat coagulation test for albumin.
• Diseases like Alzheimer's and prion diseases can occur from misfolded proteins.

Protein Purification

• Protein purification involves the isolation of specific proteins.
• This includes its importance and techniques.

Importance

• Protein purification is important in both science and medicine.

Techniques

• Techniques include salting-in and salting-out, dialysis, chromatography, and SDS-PAGE.
• Solubility is directly affected by the salt concentration of the solution.
• At low salt concentrations (salting-in), salts stabilize charged groups on a protein, attracting water. These interactions enhance protein solubility.
• At high salt concentrations (salting-out), salts trap water molecules, reducing their availability. This results in protein precipitation.

Dialysis

• Dialysis separates proteins from small solutes.
• It uses a semi-permeable membrane, where the protein solution is placed in a cellophane bag, which is immersed in a solution.
• The pores in the membrane allow water and small molecules to pass through but retain large protein molecules.
• Most dialysis membranes exclude molecules larger than 3 kDa.

Chromatography- Gel Filtration

• Gel filtration separates proteins based on size.
• This is achieved using a column filled with porous gel beads.
• Smaller molecules enter the pores and move more slowly, while larger molecules cannot enter the pores and flow through faster.
• Applications include the estimation of protein molecular weight and desalting protein mixtures.

Ion Exchange Chromatography

• Separates proteins based on charge using a column with charged resin, which carries either negative (polyanionic) or positive (polycationic) groups.
• Oppositely charged proteins bind to the resin at the appropriate pH.
• Proteins are eluted by washing with a salt solution, which disrupts electrostatic interactions.
• A gradually increasing salt concentration releases weakly bound proteins first, followed by tightly bound proteins.

Plasma Proteins Overview

• Plasma proteins are key soluble components of blood plasma, making up 60-90 g/L, about 4% of the body's total protein.
• Functions include transporting substances (e.g., hormones, drugs), maintaining hemostasis (blood clotting), and providing defense against pathogens (antibodies).
• Proteins can be separated by ammonium sulfate or gel electrophoresis.
• Most (except albumin) are glycoproteins, mening they contain carbohydrate groups.

Types

• Albumin is the major protein for maintaining osmotic pressure and transport.
• Globulins include transport and immune functions (a1, α2, β, γ).
• Fibrinogen serves as a precursor to fibrin and is essential for blood clotting.

Albumin

• Normal plasma level is 3.4–4.7 g/dL
• The liver produces about 14 g/day, accounting for 25% of hepatic protein synthesis.
• Albumin is the major plasma protein (60% of total plasma protein), present in plasma (40%) and extracellular space (60%) with a half-life of ~20 days.
• It dissolves in water and coagulates with heat.
• Found in eggs, blood, milk, and cereals.
• Functions include maintaining osmotic pressure and transport for fatty acids, bilirubin, calcium, copper, magnesium, steroid hormones, vitamins, and drugs (e.g., penicillin, aspirin).

Clinical Conditions

Hypoalbuminemia (Low Levels)

• Causes: Cirrhosis (liver disease), malnutrition, nephrotic syndrome, burns, malabsorption.
• Effect: Edema (fluid accumulation).

Hyperalbuminemia (High Levels)

• Cause: Fluid depletion (hemoconcentration).

Globulins

• Normal plasma level is 2.7-3.2 g/dL.

Functions

• α-Globulins (a1 and a2) and β-Globulins transport lipids, hormones, and trace elements; produced in the liver.
• y-Globulins provide a protective function as antibodies; made by plasma cells and B-cells in the lymphoid system.
• Properties include insolubility in water, solubility in dilute salt solutions, coagulation by heat, and precipitation at half saturation of ammonium sulfate.
• Sources include egg, blood (serum globulin), milk (lactoglobulin), muscles (myoglobin), and plants (nuts, peas).

Collagen

• Makes up 30% of total body protein and is an insoluble protein found in connective tissues like skin, bones, and tendons.
• Composition: rich in glycine (33%), proline, and lysine.
• Contains hydroxyproline and hydroxylysine formed by post-translational modifications.
• Vitamin C deficiency weakens collagen by preventing hydroxylation, reducing hydrogen bonding.
• Structure: forms a triple helix (three polypeptide chains are wound together).
• Has a firm structure due to short helical turns (3 amino acids per turn), glycine allowing tight packing, assembly into fibers, and chemical cross-linking stabilized by hydrogen bonds and hydroxyproline.

Elastin

• Elastin is a rubber-like protein that can stretch several times its length and return to its original shape when relaxed.
• It can be found in the lungs, walls of large blood vessels, and elastic ligaments.
• Composed of four interconnected polypeptide chains forming a cyclic structure called Desmosine, responsible for elastin's elasticity.
• 80% hydrophobic amino acids do not form hydrogen bonds, facilitating the elastin core's ability to stretch and recoil which provides flexibility to tissues.

Explore the effects and causes of protein denaturation, including physical and biological impacts. Learn about plasma proteins, their functions, and their significance in blood clotting and overall health. Understand conditions like hypoalbuminemia and the role of albumin in the body.