BIOCHEM LAB 3 Protein Denaturation

Biological Catalysts and Transporters

Functions of CHON

1°

Single Peptide Bonds

2°

Alpha-helix; Beta-pleated sheet

3°

Combination of 2°

4°

Combination of 3°

Denaturation

unfolding of proteins (2°,3°, 4° structure)

By heat(atoms vibrate), By strong acid(split salt linkages), by strong solvents(disrupt H bonds), By heavy metal precipitation(combine with free carboxylate anions), By organic acids(Combine with protonated amino groups)

5 ways to denature a protein

Protonated

precipitation of alkaloids

Biuret Test

Test for positive 2 peptide bonds (+Violet/-Blue)

Lead Acetate Test

test for positive sulfur in AA’s (Cysteine and Methionine;Orange + for sulfur/White - for sulfur)

Ninhydrine Test

General Test for CHON

Xanthroproetic

positive benzene in AA

Millon’s test

positive phenol in AA

Hopkin’s Cole Test

positive for Tryptophan

Sakagichi Test

positive for arginine(guanidine group)

Coagulation and Precipitation

2 ways to do denaturation test

Coagulation

a process that involves colloidal particles coming together to form larger particles that eventually settle as residue or float on the surface

defined as the process of adding chemicals to effluents to remove suspended solid wastes by neutralizing the repulsive electrical charges on particles, allowing them to aggregate into larger floccules for easier gravitational settling.

Precipitation

is the formation of a solid in a solution during a chemical reaction. When the reaction occurs, the solid formed is called the precipitate, and the liquid remaining above the solid is called the supernate

"sedimentation of a solid material (a precipitate) from a liquid solution"

form into insoluble substance

20%

percentage of albumin if concentrated

Heat, Strong Acid, Alcohol

3 ways to do coagulation denaturation test

Heavy metals, Organic Acids

2 ways to do precipitation denaturation test

Picric and Tannic Acid

2 organic acids used for precipitation denaturation test

2.5ml egg white + 47.5 ml DH2O

Contents of 5% albumin solution

globular protein

protein albumin contains:

HCL and H2SO4

soluble to 5% albumin, it’s acids disrupts hydrogen bonds and disulfide bonds

denatures and decreases solubility

what does ethanol do to proteins

blue(-)/violet(+)

color determinators for color test for AA and CHON

Yellow

Color for picric acid

Brown

Color for Tannic acid

Copper(II) sulfate and Sodium Hydroxide (and sometimes sodium potassium tartarate)

2 or 3 Biuret's reagent components

Copper(II) sulfate

Active components that reacts with peptide bonds in proteins

Sodium Hydroxide

Provides alkaline environment necessary for the reaction

Sodium potassium tartarate

Stabilizes copper ions and prevents precipitation