Factors that affect enzyme action

Experimental Purposes

1. Mastering the effects of temperature on enzyme activity; 2. Mastering the most appropriate pH of enzyme activity and mastering a method of detecting the most suitable pH; and 3. Mastering the effects of activators and inhibitors on enzyme activity."
equipment
1. Equipmenttest tube
and test tube rack,
Berries
,
ringing
water bath, ice cubes, white porcelain plate2.Reagents(1) 0.5% starch solution, 0.2% starch liquid(2) 0.3% NaCl solution, 0.85% NaCl solution.(3) Configuration of different pH buffers: (1)1/15mol/LKH
2
PO
4
liquid: pure KH
2
PO
4
9.078g/>, dissolved and diluted to 1000ml with distilled water. (2) 1/15mol/L Na
2
HPO
4
liquid: called pure Na
2
HPO
4
.2H
2
O11.815g/>, distilled water dissolved and diluted to 1000ml.
The two fluids are mixed evenly in proportion to the table below the table, and each pH buffer is 2000 pH.






(4) potassium iodide-iodine solution: 20g/> potassium iodide and 10g/> in 100ml water. Dilute 10 times before use.
(5) 1%CuSO
4
solution.
(6) 1%Na
2
SO
4
solution.
The effect of

(i) temperature on enzyme activity
because enzymes are biocatalysts, the essence of which is
protein
. Therefore, temperature sensitivity is an important characteristic of enzymes, and temperature has a double effect on the speed of enzymatic reaction.



ase as a biocatalyst, and the general catalyst shows the same temperature effect, on the one hand, the increase of temperature can increase the number of active molecules, improve the speed of enzymatic reaction, on the other hand, when the temperature exceeds a certain range, it will accelerate the denaturation rate of enzyme protein, increase the thermal denaturation of enzymes. Therefore, in the lower temperature range, the increased temperature increases the number of active molecules, the enzyme reaction speed increases with the temperature, the effect of accelerating the reaction speed is dominant, but after a certain temperature, the temperature continues to rise, the denaturation of the enzyme inactive play a leading role, the reaction speed decreases. The temperature at which the enzyme reacts at maximum speed is called the most suitable temperature of the enzyme. At the right temperature, the enzyme reacts most quickly. The most suitable temperature of an enzyme is not a constant, it is related to the length of the action time.



temperature of most animal enzymes is 37 to 40 degrees C, and the maximum temperature of plant enzymes is 50 to 60 degrees C. The stability of enzymes to temperature is related to their form of existence. The
dheating
preparation of some enzymes, although
heated
to 100 degrees C/>, did not significantly change its activity, but quickly lost its activity completely in a solution of 100 degrees C/>. Low temperatures can reduce or inhibit the activity of enzymes, but they do not inactive.



experiment, saliva amylase was used as an example. At different temperatures, the degree to which starch is hydrolyzed by salivary amylase can be determined by the color of the hydrolyzed mixture in iodine. Starch is blue with iodine. According to the size of its molecules, iodine may be blue, purple, dark brown or red. The simplest paste is iodine does not show color, maltose meets iodine does not show color.


starch -------------→dexodext-----------→ maltoseencounter iodine: blue --------→seven to red--------→ iodine1. Prepare saliva. Rinse your mouth with distilled water to remove food residue and add a little distilled water to stimulate saliva secretion. Take 1 small funnel, pad small pieces of thin skimmed cotton, directly spit into the small funnel, get filter fluid, add water to 9 ml, mix and spare.

2. Take 3 small test tubes, number, add various reagents according to the table.






3. After mixing, the 1st, 2nd and 3rd test tubes were placed in boiling water bath, warm water bath (37 to 40 degrees C/>) and ice bath 5min, add 3 drops of saliva to each tube, and continue to place 5min in the raw water bath.


4. Each tube takes out 2 drops of test fluid on the white porcelain plate, observes the color and records.

5. Then put 1, 3 tubes in 37 degrees C / > bath 5 min, each tube iodized liquid, observe color changes and analysis.




(ii) pH , activators, inhibitors on enzyme activity


environmental pH directly affects the enzyme and substrate desorption state, thus affecting the enzyme and substrate binding speed and speed. At a certain pH, the enzyme activity is greatest, and this pH becomes the most suitable pH for the enzyme. When the ambient pH is higher or lower than the most suitable pH, the activity of the enzyme decreases, and when the most suitable pH is kept away, it can even lead to the denaturation of the enzyme. Different enzymes are most suitable for pH. The most suitable pH of an enzyme is not a characteristic physical constant, and for an enzyme, the most suitable pH varies depending on the nature of the buffer and substrate.




pH for most enzymes in the human body is around 7.0. For example, the most suitable pH for salivary amylase is 6.8.










/>/>/>/>/>/>



The enzyme from inactive to active, or increase the activity of the enzyme substance called enzyme activator, the activator is mostly metal ions, such as Mg
2 plus
, K
plus
, Mn
2 plus
, and a few are anions, such as Cl
-
, there are many
organic
compound activators, such as bile salts. Any substance that inhibits the enzyme-promoting reaction is collectively called an inhibitor. A small amount of activators and inhibitors can affect the activity of enzymes, and are often specific, such as Cl
-
, which activates the activity of salivary amylase enzymes, and Cu
2 plus
, which inhibits them. However, the activators and inhibitors are not absolute, and changes in concentration may turn the activators into inhibitors.





Taking salivary amylase as an example, this experiment observed the effect of pH, activator and inhibitor on enzyme activity, and observed the method of starch hydrolyzing.




take 7 test tubes, number, add reagent


The






after mixing the tubes, and at the same time placed in a 37 to 40 degrees C/ > bath to keep warm. After about 1min, 1 drop of iodine reaction is taken out from the 3rd tube on a white porcelain plate, the color is observed, brown (if blue, the iodine reaction is still carried out once every 1min interval). Remove each tube, each iodized liquid 2 drops, shake well, observe and explain the results.






The
temperature





1. What is the temperature of salivary amylase?




2. What is the effect of low temperature on enzymes?




3. In the second experiment, why use the 3rd tube as an iodine reaction to observe the color? What's the point?




4 In the second experiment, why do you want to do a
of 4
, Na
2
SO
4
in the second experiment?