1.10 Protein desnaturalitation

Introduction

Desnaturalitation es a process in wich proteins or nucleicacids lose the quaternary, terciary and secondary structure that is present in their native State. Denaturatiom is the result of the application ofsome external stress (heat and pH change) or compounds such as a strong acid or base, a concentrated inorganic salt or organic solved.

Materials
-2x250mL beaker
-4 test tube
-Test tube rack
-10mL pipet
-Knife
-Glass marking pen
-Potato
-Distilled water
-Hydrogen perioxide
-NaCl
-HCl

Objectives

1-Study the relation between the structure and the function of proteins.                                                     
2-Understand how temperature, pH and salinity affect to the protein structure.

Catalasa activity

Caalasa is a common enzyme found in nearly all-living organisms exposed to oxygen. It catalyzes the decomposition of hidrogen perioxide (H2O2) to water an oxygen. It is a very important enzyme in protecting the cell from oxidative damage and preventing the accumulation of hidrogen perioxide.

2 H2O2 ----> 2H2O + O2

Procedure

In this experiment we are going to test the catalase activity in diferent environment situations. We  are going to measure the rate of enzyme activity under various conditions, such as different pH values and temperatures. We will measure catalase activity by observin the oxygen gas bubbles when H2O2 is destroyed. If lots of bubbles are producted, it means the reaction is happening quickly and the catalase enzyme is very active.

1-Prepare 30mL of H2O2 10% in a beaker (use a pipet).

2-Prepare 30mL of HCl 10% in a beaker.      à Solutions

3- Prepare 30mL of NaCl 50% in a beaker.

4-Peel a fresh potato tuber and cut the tissure in five cubesof 1cm3. Weigh them and equal the mass.

5-Label 5 test tubes (1, 2, 3, 4, 5)

6- Immerse 10 minutes your piece of potato inside the HCL beaker 

7- Immerse 10 minutes another piece of potato inside NaCL beaker.

8- Boil another piece of potato.

9- With a mortar, mash up the third piece of potato.

10- Prepare 5 test tubes as indicated below:

TUBE	TREATMENT
1	Raw potato
2	Boiled potato
3	Potato with HCl treatment
4	Potato with Nacl tratment
5	Mashed up potato

11- Add  5ml H2O2 in each test tube

12- With a glass-marking pen mark the heigh of the bubbles. Measure it with a ruler.

13- Compare the results of the 5 test tubes.

Table with the important parts of this experiment:

Parts:	In this experiment this was...
Independent variable	Treatment
Dependent variable	Bubble’s height
Experimental group(s)	2, 3, 4, 5 (test tube)
Control groups	1 (test tube)
Constant	Concentration of distilled water, the weight of the potato and oxigeneted water

Results

Observations

-We had to put more oxygenated water since almost no reaction occurs.

-The Mashed potato produced more activity.
-The boiled potato and tehe potato with HCl treatment produced less activity.

Questions

1.How did the temperature of the potato effect the activity of catalase? Temperature

desnaturated the catalase.

2.How did the change of the pH of the potato effects the activity of catalase? The change of the pH denaturate the catalase.

3.In wich potato treatment was catalase the most active? Why do you think this was? Mashed potato, because we had broken the cells and the catalase was more quikly.

4.An experiment was performed to test the effect of temperature and pH on the activity of Enzyme X. The following data was collected during the experiment:
a) What is the optimun pH of enzyme X? 8 (maximum activity).
b) What is the optimun temperature of enzyme X? 20 (maximum activity). c) Why do you tink enzyme X has low activity at a pH of 10? Because has been denatured.
d) Enzyme X performs critical life functions. Use the data above to explain why a fever of 40 degrees may be dangerous. Bacause is more lower than the maximum activity.

1.9 Protein identification

Introduction

Biuret's test is a chemical test used for detecting the presence of peptide bonds. Polypeptides as proteins, are chains of amino acids link together by peptide bonds.

A peptide bond can be broken by hydrolysis (the adding water). In organisms, protein molecules called enzymes facilitate the process.

Materials
-7x250mL beaker
-6 test tube
-Test tube rack
-6x10mL pipet

-Mortar
-Glass marking pen
-Gloves
-Goggles
-Milk
-Rice milk
-Egg
-Potato
-Distilleted water
-NaOH 20%
-10 drops of CuSO4 

Objectives

1. Identify peptide bonds.

2.Compare protein concentration in different foods.

Procedure

Firs of all we are going to dilute the protein

1. Add 100ml of distilled water to each 250ml beaker. Lebel them with M(milk), EW (egg white), EY (yolk), P (potato) and RM (rice milk)

2. Separate the egg white and the yolk in another beaker.

3. Smash the potato in a mortar and add some amount of the smashed potato to the P beaker.

Prepare the samples

4. Add 10ml of a dispersion of each food (M, RM, EY, EW and P) to the indicate beaker. Calculate the final concentration. All the groups will use the same dispersion from the beakers.

5. Prepare 6 test tubes (cleand and dry) and lebel (M, RM, EY, EW and P). Add 2ml of the every food dilution of each beaker.

6. Add 2ml of 20% NaOH dissolution. 

7. Shake gently and add 5 drops of CuSO4 in each tube. Allow the mixture 5 minutes. 

8. Note any colout change. Remember that proteins will turn solution pink or purple. 

9. Compare the test tubes.