enzyme kinetics
Experiment 4: Enzyme Kinetics.
Results/Discussion
Week 1 Part A:
Table 1. Enzyme activity for each assay of 4-nitroaniline formation.
Rate of 4-nitroaniline formation
Name of trial
Abs/sec
Abs/min
M/min
mol/min µmol/min #1
0.00003
0.0018
2.05x10-7
2.15 x10-10
2.15 x10-4
# 2
0.00010
0.0060
6.81x10-7
7.15x10-10
7.15x10-4
# 3
0.00020
0.0120
1.36x10-6
1.43x10-9
1.43x10-3
# 4
0.00030
0.0180
2.00x10-6
2.10x10-9
2.10x10-3
# 5
0.00040
0.0240
2.70x10-6
2.84x10-9
2.84x10-3
# 6
0.00050
0.0300
3.41x10-6
3.85x10-9
3.85x10-3
Sample Calculations
1. Changing from Abs/ sec to Abs/ mins
0.0003 Abs/ sec ( 60 sec / 1 minute ) = 0.018 Abs/ min 2. Changing from Abs/ min to M/ min (Beers-Lambert Law) C= A / El = 0.018 Abs/ min / (8800 M-1 cm -1) (1 cm) = 2.0 x 10-6 M/ min 3. Changing from M/min to mols /min = M/ min x Vol of cuvette = 2.00 x 10-6 M/ min (1050 ul / 1.0 x 106 L ) = 2.10 x 10-9 Mols/min 4. Changing from Mols/ min to umol/ min = 2.10x 10-9 Mols/min x (1.0 x 10 6 )= 2.1x10-3 umol/ min
Table 2. Average catalytic rates in abs/min.
Intervals
Average Catalytic rate (abs/min) 0 to 10 minutes
0.159
0 to 30 seconds
0.268
30 seconds to 1 minute
0.312
1 minute to 2 minute
0.255
2 minute to 3 minute
0.227
3 minutes to 5 minutes
0.177
5 minutes to 7.5 minutes
0.116
7.5 minutes to 10 minutes
0.071
Sample Calculations:
0 to 10mins:
Average Catalytic rate: 1.704-0.111abs/10-0min = 0.159 abs/min
As the table above shows, the average catalytic rates for each interval are not the same. The reaction occurs every second which means that, by the time that the enzyme was added to the substrate, it starts converting to product. Thus, when the interval takes longer (10s, 15 s and 30 s) the substrate in the reaction will get
Results/Discussion
Week 1 Part A:
Table 1. Enzyme activity for each assay of 4-nitroaniline formation.
Rate of 4-nitroaniline formation
Name of trial
Abs/sec
Abs/min
M/min
mol/min µmol/min #1
0.00003
0.0018
2.05x10-7
2.15 x10-10
2.15 x10-4
# 2
0.00010
0.0060
6.81x10-7
7.15x10-10
7.15x10-4
# 3
0.00020
0.0120
1.36x10-6
1.43x10-9
1.43x10-3
# 4
0.00030
0.0180
2.00x10-6
2.10x10-9
2.10x10-3
# 5
0.00040
0.0240
2.70x10-6
2.84x10-9
2.84x10-3
# 6
0.00050
0.0300
3.41x10-6
3.85x10-9
3.85x10-3
Sample Calculations
1. Changing from Abs/ sec to Abs/ mins
0.0003 Abs/ sec ( 60 sec / 1 minute ) = 0.018 Abs/ min 2. Changing from Abs/ min to M/ min (Beers-Lambert Law) C= A / El = 0.018 Abs/ min / (8800 M-1 cm -1) (1 cm) = 2.0 x 10-6 M/ min 3. Changing from M/min to mols /min = M/ min x Vol of cuvette = 2.00 x 10-6 M/ min (1050 ul / 1.0 x 106 L ) = 2.10 x 10-9 Mols/min 4. Changing from Mols/ min to umol/ min = 2.10x 10-9 Mols/min x (1.0 x 10 6 )= 2.1x10-3 umol/ min
Table 2. Average catalytic rates in abs/min.
Intervals
Average Catalytic rate (abs/min) 0 to 10 minutes
0.159
0 to 30 seconds
0.268
30 seconds to 1 minute
0.312
1 minute to 2 minute
0.255
2 minute to 3 minute
0.227
3 minutes to 5 minutes
0.177
5 minutes to 7.5 minutes
0.116
7.5 minutes to 10 minutes
0.071
Sample Calculations:
0 to 10mins:
Average Catalytic rate: 1.704-0.111abs/10-0min = 0.159 abs/min
As the table above shows, the average catalytic rates for each interval are not the same. The reaction occurs every second which means that, by the time that the enzyme was added to the substrate, it starts converting to product. Thus, when the interval takes longer (10s, 15 s and 30 s) the substrate in the reaction will get