Laboratory Exercise 3: Proteins and Denaturing Agents

Question 1

polymers of amino acids

Answer 1

proteins

Question 2

A typical protein may be composed of __ of amino acids

Answer 2

hundreds

Question 3

The R groups of the amino acid may be __

Answer 3

• nonpolar
• polar
• positively charged
• negatively charged

Question 4

structure of proteins that is the sequence of amino acids

Answer 4

primary structure

Question 5

structures of proteins that define the protein’s folded state

Answer 5

secondary and tertiary structures

Question 6

the state in which the protein is most active and functional. What structures and what is it called?

Answer 6

• secondary and tertiary structures
• native conformation

Question 7

Proteins are held in their native conformations by a combination of forces:

Answer 7

• hydrogen bonds
• ionic interactions
• disulfide bridges
• hydrophobic interactions

Question 8

Changing the conformation of a protein either temporarily or permanently by disrupting these forces is called __.

Answer 8

denaturation

Question 9

results in a loss of protein activity

Answer 9

denaturation

Question 10

Since the native conformation is usually the most __, disrupting the secondary and tertiary structures causes changes in __. It frequently results in the formation of a __ in the solution.

Answer 10

• water-soluble
• solubility
• solid

Question 11

Reagents or conditions that can cause denaturation are called __; these include:

Answer 11

• denaturing agents
  = heat
  = pH changes
  = alcohol
  = heavy metal salts

Question 12

describe the figure of hydrogen bonds in the lab sheet

Answer 12

N-H — O=C
O=C — H-N

(hydrogen atom that is covalently bonded to more electronegative atom oxygen–donor electron)

Question 13

describe the figure of ionic bonds in the lab sheet

Answer 13

NH2(+) (-)OOC

(formed between two ions of opposite charges, typically a metal cation and a non-metal anion, through the transfer of electrons)

Question 14

describe the figure of the disulfide bridge in the lab sheet

Answer 14

S–S

(covalent bond formed between the sulfur atoms of two cysteine amino acids in a protein)

Question 15

describe the figure of the hydrophobic interaction in the lab sheet

Answer 15

–Ar
Ar–

(nonpolar interaction between molecules in aqueous environments, where hydrophobic (water-repelling) portions of molecules tend to aggregate to minimize their contact with water molecules)

Question 16

supply kinetic energy to protein molecules, causing their atoms to vibrate more rapidly.

Answer 16

heat

Question 17

what forces can heat disrupt?

Answer 17

relatively weak forces such as hydrogen bonds and hydrophobic interactions.

Question 18

most common example is observed (heat)

Answer 18

cooking an egg

Question 19

also used in sterilization to denature and hence destroy the enzymes in bacteria.

Answer 19

heat

Question 20

• can cause a protein to denature.
• The R-groups in the amino acid chain are often charged and can form ionic bonds with a group of opposite charge.
• can change the charges on these positive and negative groups, disrupting ionic bonds.

Answer 20

Extremes of pH

Question 21

Some reagents, such as __, can form hydrogen bonds with protein molecules which will disrupt the hydrogen bonding within the molecule.

Answer 21

• ethanol

Question 22

A __ of alcohol can be used as a disinfectant because the alcohol functions to denature the proteins in bacteria. It is used because it will effectively penetrate the __

Answer 22

• 70% solution
• bacterial cell wall

Question 23

a __ coagulates proteins at the surface of the cell wall, forming a __ that prevents the alcohol from penetrating the cell.

Answer 23

-95% solution
- crust

Question 24

examples of salts of metal ions

Answer 24

• mercury(II)
• lead(II)
• silver

Question 25

They can form strong bonds with disulfide groups of proteins, disrupting disulfide bridges and salt linkages, and causing the protein to precipitate as insoluble metal-protein salts.

Answer 25

salts of metal ions

Question 26

Why are heavy metal salts, like those of mercury(II), lead(II), and silver, used as topical antiseptics?

Answer 26

- disrupt protein structures - precipitating proteins out of solution

Question 27

Why are most heavy metal salts toxic when taken internally?

Answer 27

Internally, they precipitate proteins in cells, leading to toxicity, as they come into contact with proteins of various cells.

Question 28

How do substances high in protein, such as egg whites and milk, act as antidotes for heavy metal poisoning?

Answer 28

Proteins in these substances readily combine with heavy metal ions, forming insoluble solids that can be safely eliminated from the body.

Question 29

What precaution is necessary when dealing with heavy metal poisoning and the use of protein antidotes?

Answer 29

The resulting insoluble matter must be promptly removed from the stomach using an emetic to prevent the gastric juices from destroying the protein and releasing the poisonous heavy metal ions.

Question 30

In exercise 3, the effect of several __ on the protein albumin will be studied. __ is a simple globular protein. It is soluble in __ and dilute salt solutions such as __

Answer 30

- denaturing agents - Albumin - water - isotonic saline (0.9% NaCl).

Question 31

Materials in Exercise 3: Proteins and denaturing agents

Answer 31

- 20 mL test tubes (7) - pH test strips (4) - Test tube holder (1) Alcohol lamp-tripod-wire gauze set (1) - Stirring rod (2) - 250 mL beaker (2) - Watch glass (1) - Pipet 5 mL (1) - Aspirator (1) - Wash bottle (1) - Test tube brush (1)

Question 32

Chemicals in Exercise 3: Proteins and denaturing agents

Answer 32

2 N HCl 2.5 N NaOH 0.2 M AgNO3 (Silver nitrate) (with dropper) 10% trichloroacetic acid 95% Ethanol

Question 33

Procedure in Exercise 3: Proteins and denaturing agents (1-3)```

Answer 33

1. Label seven (7) test tubes as A, B1, B2, C, D, E, and Control. 2. Prepare a stock albumen solution by adding the white of one (1) egg to 100 mL of distilled water in a beaker. Stir until the albumen is in solution. 3. Place 5 mL of the albumen solution into each of seven (7) test tubes.

Question 34

Treatment for Control Test Tube

Answer 34

Let stand at room temperature. Measure the temperature. Determine the pH with pH paper.

Question 35

Treatment for Test Tube A

Answer 35

EFFECT OF HEAT: Heat tube A in a hot water bath for a few minutes.

Question 36

Treatment for Test Tube B

Answer 36

The effect of pH changes: * To tube B1, add 2 mL of 2 N HCl. Record the pH using the pH test strips. * To tube B2, add 2 mL of 2.5 N NaOH. Record the pH using the pH test strips.

Question 37

Treatment for Test Tube C

Answer 37

The effect of metal ions: Add 5-10 drops of AgNO3 (Silver nitrate)

Question 38

Treatment for Test Tube D

Answer 38

The effect of an alkaloidal reagent: To tube D, add 1 mL of 10% trichloroacetic acid.

Question 39

Treatment for Test Tube E

Answer 39

The effects of ethanol: To tube E, add 1 mL of 95% ethanol.