Activity 19.

1 marking scheme
Investigating the primary structure of ribonuclease

Part A Comparing ribonuclease A sequences in three mammals

1 horse – 128
common minke whale – 124
red kangaroo – 122
2 K lysine
E glutamic acid
T threonine
A alanine
A alanine
A alanine
K lysine
F phenylalanine
E glutamic acid
R arginine
Q glutamine
H histidine
3

Position 1 2 3 4 5 6 7 8 9 10 11 12

Fig. 2.18 K E T A A A K F E R Q H

horse K E S P A M K F E R Q H

Position 1 2 3 4 5 6 7 8 9 10 11 12

Fig. 2.18 K E T A A A K F E R Q H

minke R E S P A M K F Q R Q H
whale

Position 1 2 3 4 5 6 7 8 9 10 11 12

Fig. 2.18 K E T A A A K F E R Q H

red E T P A E K F Q R Q H M
kangaroo

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4 The sequence of amino acids in horse pancreatic ribonuclease compared with bovine
pancreatic ribonuclease:
• nine of the twelve positions have the same amino acids
• the amino acids in positions 1, 2, 5 and 7–12 are the same (K, E, A and KFERQH)
• the amino acids in positions 3, 4 and 6 are different (T–S, A–P and A–M).
The sequence of amino acids in minke whale pancreatic ribonuclease compared with bovine
pancreatic ribonuclease:
• seven of the twelve positions have the same amino acids
• the amino acids in positions 2, 5, 7, 8 and 10 to 12 are the same (E, A, K, F and R, Q and H)
• the amino acids in positions 3, 4, 6 and 9 are different (K–R, T–S, A–P, A–M and E–Q).
The sequence of amino acids in red kangaroo pancreatic ribonuclease compared with bovine
pancreatic ribonuclease:
• only one of the twelve positions has the same amino acid
• A is in position 4 in both molecules
5 If the sequence for red kangaroo is moved along one position, there is a greater degree of
similarity between the two.

Position 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Fig. 2.18 K E T A A A K F E R Q H M D S

red E T P A E K F Q R Q H M D T E
kangaroo

Position 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Fig. 2.18 K E T A A A K F E R Q H M D S

red E T P A E K F Q R Q H M D T
kangaroo

This now shows that there are ten positions that are occupied by the same amino acids.
This suggests that the gene for pancreatic ribonuclease in the red kangaroo has one less triplet than
the genes for this enzyme in the other three mammals.
6 Tryptophan, W or Trp. It is found at position 39 in the horse ribonuclease.
No. Tryptophan is not in bovine ribonuclease.
7 Position 39 in bovine ribonuclease and minke whale ribonuclease is occupied by arginine (R or
Arg). The DNA triplets for this amino acid are GC_ (where _ represents any base, A, C, T or G),
TCT and TCC.
There is only one DNA triplet for tryptophan – ACC.
It is possible for the triplets TCC and GCC to change by a substitution mutation to ACC. This
would replace arginine by tryptophan in the amino acid sequence.
8 Cysteine (C). There are eight cysteine residues and, therefore, four disulfide bonds. They are at
positions: 26 – 84, 40 – 95, 58 – 110, 65 – 73

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9 In the horse and minke whale, the disulphide bonds are all in exactly the same positions. In the red
kangaroo, they are in one place to the left. This corresponds to the idea that red kangaroo
ribonuclease has one less amino acid at the beginning of the sequence.
10 These regions may be active sites of enzymes or binding sites of cell surface receptors that have
the same shape in order to function efficiently whatever species is the source of the proteins
concerned. The shapes of active sites and binding sites are determined by the R groups of the
amino acids that make up these regions.
11 Possible suggestions include the following.
• Investigating the roles of proteins in genetic diseases such as haemoglobin in sickle cell
 anaemia (pages 388 of the Coursebook) and CFTR in cystic fibrosis (pages 478–480
 of the Coursebook).
• Locating and studying key regions of proteins, such as active sites of enzymes and
 antigen-binding sites of antibodies.
• Studying the interaction between natural signalling molecules and cell surface receptors
 (pages 77–79 and pages 226–237 of the Coursebook).
• Studying the interaction between drugs and proteins, such as cell surface receptors. Using
 this information in the design of new drugs.
• Providing molecular evidence for evolution (pages 404–408 of the Coursebook).

Part B Using online tools to investigate the structure of ribonuclease A (RNase A)

1 This is the full sequence for bovine pancreatic ribonuclease:

1 10 20 30 40 50 60
MALKSLVLLS LLVLVLLLVR VQPSLGKETA AAKFERQHMD SSTSAASSSN YCNQMMKSRN

70 80 90 100 110 120

LTKDRCKPVN TFVHESLADV QAVCSQKNVA CKNGQTNCYQ SYSTMSITDC RETGSSKYPN

130 140 150

CAYKTTQANK HIIVACEGNP YVPVHFDASV

This sequence has 150 amino acids rather than 124 as shown in Figure 2.18 in the Coursebook.
This is because the full sequence has an additional 26 amino acids at the start of the sequence
(from position 27 on, the sequence is the same). This initial sequence of 26 amino acids is a signal
sequence or signal peptide at the N terminal of the protein. Signal peptides are present at the N
terminal of proteins that are secreted from cells. The cell uses the signal sequence to make sure the
protein passes through the endoplasmic reticulum and the Golgi body to enter a vesicle that
releases the protein by exocytosis. Ribonuclease is secreted from pancreatic cells into the
pancreatic duct and from there it passes into the small intestine.

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