CHEM 271 Chapter Notes - Chapter Tutorial 4: Azide, Central Cross-Island Highway, James Joyce

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kzala5602

1 Dec 2021

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Concordia University

Department

Chemistry

Course

CHEM 271

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Enzyme lab: virtual enzyme kinetics qwertyuiopasdfghjklzxcvbnmqwertyui opasdfghjklzxcvbnmqwertyuiopasdfgh jklzxcvbnmqwertyuiopasdfghjklzxcvb nmqwertyuiopasdfghjklzxcvbnmqwer tyuiopasdfghjklzxcvbnmqwertyuiopas dfghjklzxcvbnmqwertyuiopasdfghjklzx cvbnmqwertyuiopasdfghjklzxcvbnmq wertyuiopasdfghjklzxcvbnmqwertyuio pasdfghjklzxcvbnmqwertyuiopasdfghj klzxcvbnmqwertyuiopasdfghjklzxcvbn mqwertyuiopasdfghjklzxcvbnmqwerty uiopasdfghjklzxcvbnmqwertyuiopasdf ghjklzxcvbnmqwertyuiopasdfghjklzxc vbnmqwertyuiopasdfghjklzxcvbnmrty uiopasdfghjklzxcvbnmqwertyuiopasdf ghjklzxcvbnmqwertyuiopasdfghjklzxc vbnmqwertyuiopasdfghjklzxcvbnmqw. Precision measurement under constant condition(s=1. 73 nm, i=1, ph=7. 5 ) 2. 8 mole product per min per mg enzyme. I calculated the estimated km value by using the following formula; The following graph shows that the optimum ph of protease is 7. 5 because the rate of reaction is at its higher here. 2 (michaelis-menten plot for enzyme pr/49-140-111 without inhibitor to determine km and vmax) Determination of km and vmax without inhibitor at optimum ph 7. 5. Determination of km and vmax with inhibitor at optimum ph 7. 5. 3 (michaelis-menten plot for enzyme pr/49-140-111 with inhibitor to determine km and vmax) The effect of ph on the km and vmax is that whenever ph increases, km decreases. The enzyme protease has an optimum ph of 7. 5 after testing with different ph levels ranging from 5 to 9.

Related Questions

Determine

1. The alpha value, A420/ (umole ONPG/mL)

2. The specific activity

A420/ minutes * (umole ONP/ A420 change)/ protein (mg) = unit/ mg

3. Use the michaelis-menten equation to determine, Vmax and Km values

using the following Data provided

E= 50 ug/mL	C1= 4 mg/mL
Time (sec)	A420
30	0.92
60	1.8
120	3.63
180	4.58
300	7.16
600	10.86
1800	12.34

where

E= the enzyme concentration, ug/ML

C= the substrate ONPG concentration, mg/mL

Time = incubation time, second

A420= abosorbance at 420nm

Molecular weight of ONPG 301.3 g/mole

An experimenter is working with an enzyme that, on a per milligram basis, produces a certain number of Âµmol of product per minute at optimal temperature and pH. The product of the enzyme-catalyzed reaction is a desirable fine chemical, and thus the researcher designs a pilot biosynthetic reaction to produce a target number of Âµmol of the desired product. The experimenter starts the reaction by adding 1.0 milligram of the enzyme. The rate of product formation reveals that the enzyme activity is normal. (The researcher is able to continually remove the product as it is produced.) To the astonishment of the experimenter, an assistant inexplicably adds some sodium hydroxide to the reaction after 1 min. (The assistant misunderstood the protocol.) Thirty seconds after the reaction was stopped by the addition of the NaOH, the experimenter adds additional buffer and is pleased to see that the reaction is proceeding again, but at three-quarters (3/4) of its normal activity. Furthermore, it takes 3Â½ minutes longer to produce the target amount of product than it should have. Had the enzyme been allowed to produce an additional 90 Âµmol of product prior to the addition of base, and then, 30 seconds later, the addition of buffer, it would have taken 3 minutes longer to produce the target number of Âµmol of product. What is the specific activity of the enzyme (Âµmol of product per minute per milligram of enzyme)? Enter your answer to the nearest ones.

Given the information in Part A, what was the target amount of product (in Âµmol) that was desired? Enter your answer to the nearest ones.

ceruleancricket955

Please check my work. I am have to bring my grade up so I need to make sure I get these right this week Here are my notes and answers from lab this week please let me know if I missed any.

NOTES:

Experiment 1: The Effect of Temperature on Catalase Activity

Test tube #1(10ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 5 mL of water

Add 1 mL of catalase

Observe bubbles: No bubbles observed (or very, very little)

Color changed to red

Test Tube #2 (21.5ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 5 mL of water

Add 1 mL of catalase

Observe bubbles: Bubbles appeared quickly and constantly for the first 10 seconds then stopped

Color changed to red

Test Tube #3 (40ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 5 mL of water

Add 1 mL of catalase

Observe bubbles:Bubbles appeared slowly and Sporadically for about 10 seconds then stopped.

Color changed to red

Test Tube #4(60ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 5 mL of water

Add 1 mL of catalase

Observe bubbles: Bubbles appeared instantly and continued sporadically continued for 10 seconds

color changed to red

Test Tube #5 (80ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 5 mL of water

Add 1 mL of catalase

Observe bubbles:Bubbles appeared instantly, and sporadically continued for about 10 seconds then stopped.

Color of solution changed to red.

Experiment 2: The Effect of Substrate Concentration on Catalase Activity

Test Tube #1

Add 8mL Water

Add 2mL 3% Hydrogen Peroxide (mL)

Add 0.1 mL of catalase

Time for bubbles to stop forming: .38 seconds

Test Tube #2

Add 5mL Water

Add 5mL 3% Hydrogen Peroxide (mL)

Add 0.1 mL of catalase

Time for bubbles to stop forming: 32 seconds

Test Tube #3

Add 2mL Water

Add 8mL 3% Hydrogen Peroxide (mL)

Add 0.1 mL of catalase

Time for bubbles to stop forming: 28 seconds

Test Tube #4

Add 0mL Water

Add 10mL 3% Hydrogen Peroxide (mL)

Add 0.1 mL of catalase

Time for bubbles to stop forming: No bubbles

Experiment 3: The Effect of pH on Catalase Activity

Test Tube #1 (21.5ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 5 mL of pH 2 buffer

Add 1 mL of Catalase Solution

Observation: No bubbles

Test Tube #2 (40ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 5 mL of pH 6 buffer

Add 1 mL of Catalase Solution

Observation: bubbles appear immediately

Test Tube #3 (60ÃÂ°C)

Add 5 mL of 3% hydrogen peroxide

Add 10mL of pH 10 buffer

Add 1 mL of Catalase Solution

Observation: no bubbles.

Experiment 1: The Effect of Temperature on Catalase Activity

Lab Results

At which temperatures did catalase function somewhat, but not optimally?

40 degrees the bubbles appeared slowly and continued sporadically for about 10 seconds before stopping. At 60 & 80 degrees the bubbles appeared immediately but were sporadic as well.

Data Analysis

Why did catalase fail to function at 80 Â°C?

Because the Enzyme catalase denatures

Experiment 2: The Effect of Substrate Concentration on Catalase Activity

Lab Results

Fill in the table below with your results from the substrate concentration experiment.

Substrate Concentration Results
Test Tube	Bubbles Formation (min)
1	00:38 seconds
2	00:32 seconds
3	00:28 seconds
4	No bubbles.

Data Analysis

Before the catalase concentration was saturated with hydrogen peroxide, why did the enzyme function vary with substrate concentration?

The enzymes had varied results before the catalyst concentration was changed due to the differences in temperature. In experiment one all measurements were the same, the variable was the temperature. In experiment two, the concentrations of each material were the variable along with the temperature.

Experiment 3: The Effect of pH on Catalase Activity

Lab Results

What was the pH value at which catalase functioned the best?

pH 6 buffer

Data Analysis

Why did catalase only function in a limited range of pH values?

Conclusions

What are the enzyme, substrate, and product in these experiments?

The enzyme is Catalase

The Substrate is Hydrogen peroxide

The product was the bubbles.

What role does catalase play in the mammalian liver, or any other cell it is found in?

Catalase is an enzyme in the liver that breaks down harmful hydrogen peroxide into oxygen and water. When this reaction occurs, oxygen gas bubbles escape and create foam.

Reference:

https://www.scientificamerican.com/article/bring-science-home-liver-helping-enzymes/

CHEM 271 Chapter Notes - Chapter Tutorial 4: Azide, Central Cross-Island Highway, James Joyce

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