Section 427 & 527

Lab 3: Enzymes

Bio 1AL

Part I: Enzyme Concentration & Activity

Data should be doubling because the Enzyme concentration is doubling. More enzymes =
more likely that E and S will collide at the right angle, speed, & orientation. This means that
it is more likely to form the ES complex and convert Substrate into Product. The
relationship should be linear (given [S] should be in excess).
Below is the most ideal set of data between my two lab sections:
50 ng/mL
100 ng/mL
200 ng/mL
400 ng/mL

0.535 uMoles Maltose

1.148 uMoles Maltose
2.011 uMoles Maltose
2.973 uMoles Maltose
It should look mostly linear.

Enzyme Concentration & Activity

3.500
uMoles Maltose

3.000
2.500
2.000
1.500
1.000
0.500
0.000
50 ng/ml

100 ng/ml

200 ng/ml

400 ng/ml

Amylase Concentration (Enzyme)

Part IIA: Temperature

Optimum Temperature for Amylase = 50C.
Initially you will see an increase. This is because increasing temperature results in more
kinetic energy = more collisions b/t E & S. However, at a certain temperature, the enzyme
will reach its optimum rate and begin to denature after that. (Too much energy causes the
bonds holding the protein together to break apart which changes the shape of the enzyme
-> enzyme can no longer function).
4C
21C
37C
50C
90C

0.233 uMoles Maltose

0.808 uMoles Maltose
1.684 uMoles Maltose
2.204 uMoles Maltose
0.347 uMoles Maltose
You should see an increase up until the optimum and then a decrease.

Section 427 & 527

Lab 3: Enzymes

Bio 1AL

2A: Temperature
uMoles Maltose

2.500
2.000
1.500
1.000
0.500
0.000
4

21

37

50

90

Temperature C

Part IIB: Km (Varying Substrate Concentration & Activity)

You will see 3 parts to the graph.

A) Linear at first: this is b/c as you increase the amt of substrate,

you are increasing the chance that it will collide with an enzyme and
form the ES complex.

B) Starts to curve: At this point, most of the enzymes are being used
so the rate of increase slows down.

C) Plateaus/levels off at Vmax: All of enzyme are saturated with

substrate and can't work any faster.

Think about it this way:

If you had 5 GSIs (enzymes) in the room, they can each help 1 person out at a time. If
there is only 1 student (substrate), then we are helping 1 student/time. If we increase the
amt of students to 2, then we can help 2 students/time. Once we reach 5 students, then all
of the GSIs are working so we are helping 5 students/time. If we increase the amt of
students to 6 (or more), the GSIs still can't help more than 5 students/time since there are
only 5 of us (and we can't be in two places at once)!
0.125% Starch
0.25% Starch
0.50% Starch
1.00% Starch
2.00% Starch
3.00% Starch

0.131 uMoles Maltose

0.285 uMoles Maltose
0.574 uMoles Maltose
0.906 uMoles Maltose
0.980 uMoles Maltose
1.043 uMoles Maltose

Section 427 & 527

Lab 3: Enzymes

Bio 1AL

2B: Km
1.200
uMoles Maltose

1.000
0.800
0.600
0.400
0.200
0.000
0.125

0.25

0.5

Substrate Concentration (% Starch)

Part IIC: Saliva

To find the conc of undiluted saliva, you want to first pick a data point from you table in part
6. For my example I will use (1/100 dilution, N uMoles Maltose). You will then take that N
uMoles of Maltose and find it on the Y-axis on your graph in Part I (Question 2). Next, you
want to find the corresponding X-axis value. This is the concentration of amylase in your
1/100 dilution of saliva. To find the undiluted concentration, multiply by the denominator
(100 in this case).
Ideally, the data should have been decreasing by a factor of 10 each time. This is because
we are decreasing the concentration of our saliva by a factor of 10 with each dilution.
(Similar reasoning to Part I).

1/100 Dilution
1/1000 Dilution
1/10000 Dilution
1/100000 Dilution

3.745 uMoles Maltose

2.084 uMoles Maltose
0.267 uMoles Maltose
0.092 uMoles Maltose

Section 427 & 527

Lab 3: Enzymes

Bio 1AL

uMoles Maltose

2C: Saliva
4.000
3.500
3.000
2.500
2.000
1.500
1.000
0.500
0.000
1/100

1/1000

1/10000

1/100000

Dilution

Part IID: pH
Optimum = 7.
As explained in class, each enzyme has an optimum pH at which it operates. For amylase,
this is physiological pH (pH 7) since it is found within our body. As you move away from
this optimum pH, enzyme activity quickly decreases as the enzyme becomes denatured.
4
5.5
7
8.5
10

0.454 uMoles Maltose

1.427 uMoles Maltose
1.705 uMoles Maltose
1.464 uMoles Maltose
0.145 uMoles Maltose

uMoles Maltose

pH
1.800
1.600
1.400
1.200
1.000
0.800
0.600
0.400
0.200
0.000
4

5.5

7
pH

8.5

10