Name .
Period .
Date .
Science
Cytochrome C Comparison Lab
PURPOSE: To compare the relatedness between organisms by examining the amino acid sequence in the
protein, Cytochrome C.
BACKGROUND: Genes are made of DNA and are inherited from parent to offspring. Some DNA
sequences code for mRNA which, in turn, codes for the amino acid sequence of proteins. Cytochrome C
is a protein involved in using energy in the cell. Cytochrome C is found in most, if not all, known
eukaryotes. Over time, random mutations in the DNA sequence occur. As a result, the amino acid
sequence of Cytochrome C also changes. Cells without usable Cytochrome C are unlikely to survive.
METHOD:
A . Compare the amino acid sequence of Cytochrome C in various organisms.
1. Mark the amino acids which are different Use the example to show you how.
- - - - - G D V A K G K K T F V Q K C A Q C H T V E N G G K H K V G P N L W G L F G R K T
- G V P A G D V E K G K K I F V Q R C A Q C H T V E A G G K H K V G P N L H G L F G R K T
2.
Note that some of the amino acids are always the same in all species, these have been shaded blue
or light gray.
Count and record the total number of differences.
Tuna &
Fly:
24
3.
Note: Be sure to study the entire molecule.
Each protein sequence has 103-112 amino acids; the sequence extends onto two (2) lines.
4. Share your data with the rest of the class and complete Table 1.
B . Make a branching tree, or cladogram using the data in Table 1.
s
ie
ec
sp
sp
ec
ie
s
2
The two most closely related species have the fewest differences in amino acid sequence.
Place the two most closely related species on the two shortest branches of the tree.
1
1.
3.
ie
ec
sp
sp
ec
ie
s
s
3
4
2. Place the next two closest species on the next shortest branches.
Place the species which is the next closest on the next longest branch.
Continue until all the species have been placed.
C. Answer the questions.
Cytochrome Clab pt 2
kew 04/02
Page 1 of 4
Name _________________
AMINO ACID SEQUENCES IN CYTOCHROME-C PROTEINS FROM DIFFERENT SPECIES
Amino Acid Number ---->
Human, Chimpanzee.
Rhesus monkey.....
Horse ............
Donkey ...........
Common zebra .....
Pig, Cow, Sheep...
Dog...............
Gray whale .......
Rabbit............
Kangaroo..........
Chicken, Turkey...
Penguin...........
Pekin duck........
Snapping turtle...
Rattlesnake.......
Bullfrog..........
Tuna..............
Screwworm fly.....
Silkworm moth.....
Tomato horn “worm”
Wheat ............
Rice .............
Baker's yeast (Fungus)
Candida yeast(Fungus)
Neurospora (Fungus)
1
A
A
-
2
S
S
-
3
F
F
P
-
4
S
S
T
A
-
5
G
G
G
E
E
E
P
G
6
V
V
V
A
A
F
F
F
7
P
P
P
P
P
K
E
S
8
A
A
A
P
P
A
0
A
[CONTINUED FROM ABOVE]
60
Amino Acid Number-> 0 1 2 3 4 5 6 7 8 9
Human, Chimpanzee. N K N K G I I W G E
Rhesus monkey..... N K N K G I T W G E
Horse ............ N K N K G I T W K E
Donkey ........... N K N K G I T W K E
Common zebra ..... N K N K G I T W K E
Pig, Cow, Sheep... N K N K G I T W G E
Dog............... N K N K G I T W G E
Gray whale........ N K N K G I T W G E
Rabbit............ N K N K G I T W G E
Kangaroo.......... N K N K G I I W G E
Chicken, Turkey... N K N K G I T W G E
Penguin........... N K N K G I T W G E
Pekin duck........ N K N K G I T W G E
Snapping turtle... N K N K G I T W G E
Rattlesnake....... N K N K G I I W G D
Bullfrog.......... N K N K G I T W G E
Tuna.............. N K S K G I V W N N
Screwworm fly..... N K A K G I T W Q D
Silkworm moth..... N K A K G I T W G D
Tomato horn “worm” N K A K G I T W Q D
Wheat............. N K N K A V E W E E
Rice ............. N K N M A V I W E E
Baker's yeast..... N I K K N V L W D E
Candida yeast..... N K R A G V E W A E
Neurospora (Fungus) N K Q K G I T W D E
Symbols in light blue or gray represent
(Adapted from Strahler, Arthur, Science
Cytochrome Clab pt 1
kew 04/02
9
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
10
0 1
D V
D V
D V
D V
D V
D V
D V
D V
D V
D V
D I
D I
D V
D V
D V
D V
D V
D V
N A
N A
N P
N P
S A
S A
D S
2
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
A
E
E
D
D
K
K
K
K
3
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
N
N
A
A
K
K
K
4
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
5
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
A
E
A
A
A
6
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
T
T
N
7
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
I
T
I
I
I
I
I
L
L
L
8
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
F
9
I
I
V
V
V
V
V
V
V
V
V
V
V
V
S
V
V
V
V
V
K
K
K
K
K
20
0 1
M K
M K
Q K
Q K
Q K
Q K
Q K
Q K
Q K
Q K
Q K
Q K
Q K
Q K
M K
Q K
Q K
Q R
Q R
Q R
T K
T K
T R
T R
T R
70
80
0 1 2 3 4 5 6 7 8 9 0 1 2
D T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
E T L M E Y L E N P K K Y
E T L M E Y L E N P K K Y
E T L M E Y L E N P K K Y
E T L M E Y L E N P K K Y
E T L M E Y L E N P K K Y
E T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
E T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
D T L M E Y L E N P K K Y
D T L F E Y L E N P K K Y
D T L F E Y L E N P K K Y
D T L F E Y L E N P K K Y
N T L Y D Y L L N P K K Y
N T L Y D Y L L N P K K Y
N N M S E Y L T N P K K Y
P T M S D Y L E N P K K Y
N T L F E Y L E N P K K Y
amino acids which show NO
& Earth History, 1987, p.
2
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
3
S
S
A
A
A
A
A
A
A
A
S
S
S
A
G
A
A
A
A
A
A
A
E
A
A
4
Q
Q
Q
Q
Q
Q
Q
Q
Q
Q
Q
O
Q
Q
T
Q
Q
Q
Q
Q
Q
Q
L
E
E
5
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
6
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
7
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
G
3 4 5 6 7 8
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
I P G T K M
differences
348)
8
V
V
V
V
V
V
V
V
V
V
V
V
V
V
V
C
V
V
V
V
V
V
V
I
E
9
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
E
D
D
E
E
G
30
0 1
K G
K G
K G
K G
K G
K G
K G
K G
K G
K G
K G
K G
K G
K G
E G
K G
N G
A G
A G
A G
A G
K G
K G
A G
G N
2
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
A
A
G
G
L
3
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
G
G
P
P
T
4
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Q
5
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
90
9 0 1 2 3 4 5 6
I F V G I K K K
I F V G I K K K
I F A G I K K K
I F A G I K K K
I F A G I K K K
I F A G I K K K
I F A G I K K T
I F A G I K K K
I F A G I K K K
I F A G I K K K
I F A G I K K K
I F A G I K K K
I F A G I K K K
I F A G I K K K
V F T G L K K K
I F A G I K K K
I F A G I K K K
I F A G L K K P
V F A G L K K A
V F A G L K K A
V F P G L K K P
V F P G L K K P
A F G G L K K E
A F G G L K K A
A F G G L K K D
in any organism
6
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
V
V
V
V
V
Q
Q
V
V
I
7
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
8
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
P
9
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
A
40
0 1
L H
L H
L H
L H
L H
L H
L H
L H
L H
L N
L H
L H
L H
L N
L H
L Y
L W
L H
L H
L H
L H
L N
L H
L H
L H
2
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
3
L
L
L
L
L
L
L
L
L
I
L
I
L
L
L
L
L
L
F
F
L
L
I
I
L
100
7 8 9 0 1 2 3 4
E E R A D L I A
E E R A D L I A
T E R E D L I A
T E R E D L I A
T E R E D L I A
G E R E D L I A
G E R A D L I A
G E R A D L I A
D E R A D L I A
G E R A D L I A
S E R V D L I A
S E R A D L I A
S E R A D L I A
A E R A D L I A
K E R T D L I A
G E R Q D L I A
G E R Q D L V A
N E R G D L I A
N E R A D L I A
N E R A D L I A
Q D R A D L I A
Q E R A D L I S
K D R N D L I T
K D R N D L V T
K D R N D I I T
on the list, so
4
F
F
F
F
F
F
F
F
F
F
F
F
F
I
F
I
F
F
Y
F
F
F
F
F
F
5
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
S
G
5 6
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y L
Y M
F M
you
6
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
R
7
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
K
Q
Q
H
H
K
7 8
K K
K K
K K
K K
K K
K K
K K
K K
K K
K K
K D
K D
K D
K D
K E
K S
K S
K S
K E
K Q
K K
K E
K K
L E
K E
can
8
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
S
S
S
S
T
9
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
50
0 1
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
Q A
T T
T T
Q A
Q A
S V
2
P
P
P
P
P
P
P
V
V
P
E
E
E
E
V
A
E
A
P
P
A
P
Q
Q
D
3
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
4
Y
Y
F
F
F
F
F
F
F
F
F
F
F
F
Y
F
Y
F
F
F
Y
Y
Y
Y
Y
5
S
S
T
S
S
S
S
S
S
T
S
S
S
S
S
S
S
A
S
S
S
S
S
S
A
6
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
7
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
S
S
S
S
T
T
T
8
A
A
D
D
D
D
D
D
D
D
D
D
D
E
A
D
D
N
N
N
A
T
D
D
D
9
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
110
9 0 1 2
A T N E
AMINO ACID SYMBOLS
A T N E
A = Alanine
A T N E
C = Cysteine
A T N E
D = Aspartic acid
A T N E
E = Glutamic acid
A T N E
F = Phenylalanine
A T K E
G = Glycine
A T N E
H = Histidine
A T N E
I = Isoleucine
A T N E
K = Lysine
A T S K
L = Leucine
A T S K
M = Methionine
A T A K
N = Asparagine
A T S K
P = Proline
A T A K
Q = Glutamine
A C S K
R = Arginine
A T S S = Serine
A T K T = Threonine
S T K V = Valine
A T K
W = Tryptophan
A T S S
Y = Tyrosine
A T S A C E A S K A T A ignore them.
Page
2
0 of 4
9
Name
Table 1: Differences between amino acid sequences in Cytochrome C protein for nine species.
horse
horse
donkey
whale
chicken
penguin
snake
moth
0
donkey
whale
1
0
4
4
0
chicken penguin
11
10
9
0
13
12
10
3
0
snake
moth
yeast
wheat
20
21
18
18
17
0
29
28
27
29
27
29
0
45
46
45
46
45
46
48
0
46
45
44
46
46
46
43
47
0
yeast
wheat
Cladogram, or Branching Tree,
Based on Cytochrome C Amino Acid Sequence
Cytochrome Clab pt 2
kew 04/02
Page 3 of 4
Name
QUESTIONS:
1. How many Cytochrome C amino acid sequence
differences are there between chickens and turkeys?
2. Make a branching tree, or cladogram for chickens, penguins, and turkeys.
3. a.
b.
Predict the number of Cytochrome C amino acid
sequence differences you would expect to see between i. horse and zebra
ii. donkey and zebra
What other information did you use to make this prediction?
4. Use this Cytochrome C sequence difference data to add a branch to the tree on page 3.
silkworm
horse
penguin
snake
tuna
yeast
moth
tomato
hornworm
26
18
24
28
5
42
moth
wheat
38
5. List three other things used to determine how organisms are related to each other.
6. Explain why more closely related organisms have more similar Cytochrome C.
Cytochrome Clab pt 2
kew 04/02
Page 4 of 4
7. Other data, including other genes, suggests that fungi are more closely related to animals than plants.
What are some reasons that the Cytochrome C data suggests that fungi, plants, and animals are equally
distantly related?