To Observe Enzyme Kinetics Biology Essay

Enzymes are accelerators, normally proteins, that decrease the activation energy needed for the reaction and therefore they speed up the rate of a chemical reaction. Every enzyme binds to it substrate specifically, where the enzyme binds to its substrate called the activation site. There are several factors that consequence the activity of enzymes ; concentration of substrate molecules, temperature, presence of inhibitors and pH. [ 1 ] [ 2 ]

2.2 ) Inhibitors

Substances that can cut down or halt the activity of an enzyme by barricading or falsifying the active site of the enzyme are called inhibitors. There are two types of inhibitors ; competitory or non-competitive. Competitive inhibitors binds to the active site of the enzyme so, they block it for the substrate. They compete with the substrate molecule to adhere to the active site. Non-competitive inhibitors binds to other parts of the enzyme and makes a conformational alteration in the active site of the enzyme so that the substrate ca n’t adhere to the active site because there is a lock and cardinal relation between them. [ 2 ]

2.3 ) Michaelis-Menten equation:

The michaelis menten theoretical account gives information about how an enzyme plant and the enzyme dynamicss. The equation relates the initial reaction rate to the substrate concentration. It describes the rates of irreversible reactions. [ 3 ]

Tocopherol: enzyme

Second: substrate

Einsteinium: enzyme-substrate composite

Phosphorus: merchandise

Vmax: maximal speed achieved by the system at maximal substrate concentration

Kilometers: the Michaelis invariable, substrate concentration at which the reaction speed is the half of the maximal speed.

Michaelis-menten graph is non additive. It is hard to gauge Km and Vmax accurately so there are some linerizations developed ; Lineweaver-Burke secret plan, Eadie Hofstea diagram and Hones-woolf secret plan.

3 ) EQUIPMENT AND CHEMICALS

3.1 ) equipments:

Cuvettes

Spectrophotometer

Pipet

Tips

Tubes

3.2 ) chemicals:

Sodium phosphate buffer 0,05 M ; pH 7

Mushroom tyrosinase, 100 units/ml

L-Dopa ( 20 milliliter pergroup, 7.60 ten 10-3 M )

Cinnamic acid, 50mg/100mL

4 ) Procedure

Sodium phosphate buffer, L-DOPA and cinnamic acerb solutions were prepared before the experiment.

5 trial tubings were prepared. L-DOPA was added 0.75 milliliters to all trial tubings.

Buffer was added to all trial tubings at different sums. ( buffer sums were calculculated because we took the half of the solutions that is written and tried to sum the whole solution up to 3 milliliters.

Tyrosinase was added to do the solution 3 milliliter in entire.

Optical density values were measured in every 30 seconds in 2 proceedingss.

Test tubings were prepared and this clip tyrosinase sum was kept changeless at 0.05 milliliter.

L-DOPA was added otherwise in all tubings ; 0.05, 0.1, 0.2, 0.4, 0.5, 0.75.

Buffer sum was added harmonizing to the computations to do the solution 3ml in entire.

Optical density values were measured in every 30 seconds for 2 proceedingss.

Test tubings were prepared. Tyrosinase was kept changeless at 0.05 and cinnamic acid was kept changeless at 0.4 milliliter.

L-dopa was added otherwise in all tubings ; 0.05, 0.1, 0.2, 0.4, 0.5, 0.75.

Buffer was added to the tubings to finish the solution to 3 milliliters in entire.

Optical density values were measured in every 30 seconds for 2 proceedingss.

5 ) Consequence

Sample 1:

30 seconds:

Sample

Optical density

1

0.860

2

0.890

3

0.940

4

0.940

5

0.940

60 seconds:

Sample

Optical density

1

0.820

2

0.940

3

0.950

4

0.930

5

0.940

90 seconds:

Sample

Optical density

1

0.880

2

0.920

3

0.930

4

0.930

5

0.940

120 seconds:

Sample

Optical density

1

0.890

2

0.930

3

0.940

4

0.950

5

0.970

Sample 2:

120 seconds:

Sample

Optical density

1

0.700

2

0.930

3

0.940

4

0.950

5

0.950

6

0.945

Sample 3:

120 seconds:

Sample

Optical density

1

0.205

2

0.345

3

0.500

4

0.790

5

0.820

6

0.890

6 ) Discussion:

In this experiment our intent was to detect enzyme dynamicss and to detect the alterations when an inhibitor binds to the enzyme. We used tyrosinase enzyme in this experiment. And in order to detect the dynamicss we prepared three different samples with different sums of enzymes and buffers. And in one of the samples we used an inhibitor.

In the first sample we took the L-dopa sum the same in all the tubings. And we changed the enzyme sum and buffer sum were changed. The graphs which were obtained in two proceedingss for every 30 seconds, shows us that after a certain clip the rate reaches to its bound. It ‘s the same for the 2nd sample. In this sample we used changeless sum of enzyme this clip, and changed the others. In the 3rd sample we added an inhibitor. It was a non-competitive inhibitor, cinnamic acid. Which means it decreases the velocity of the reaction. It makes a conformational alteration in the active site of the enzyme. And because of it enzymes involvement for the substrate is non altering. When we look at the graphs and compare them with and without the substrate, less sum of merchandise is formed in the same clip period.