Download Life Sciences 1a Practice Problems 6

Life Sciences 1a
Practice Problems 6
1. HMG CoA Reductase is an enzyme involved in the biosynthesis of cholesterol. The
protein and mRNA sequences were identified before the genomic sequence, and it
was determined that the mRNA is 4471 nucleotides long, and encodes a protein of
888 amino acids. During the human genome project, the genomic sequence for
HMG CoA Reductase was found on chromosome 5 spread out over 24,826
nucleotides.
a) Why is there such a big difference between the length of the HMG CoA gene
found on chromosome 5 and the length of the mRNA?
b) If all 24,826 nucleotides encoded a protein, how long would it be?
c) Based on your understanding of the nucleic acid triplet code, how many
nucleotides of mRNA sequence are necessary to encode a protein of 888 amino
acids?
d) Is there a disparity between your answer to C and the the length of HMG CoA
Reductase mRNA? If so, please explain a resolution to this difference.
e) In the bakers yeast, S. cerevisiae, it has been found that the mRNA and the
genomic sequence for HMG CoA Reductase are the exact same length. What
does this information tell you about how this gene is organized in yeast?
The Genetic Code:
2. During translation, does the peptide extend in the C-terminal or N-terminal
direction?
3. In one of the first experiments to decipher the genetic code, Marshall Nirenberg
synthesized a strand of poly(U) mRNA (that is, a strand that reads 5’ – UUU…UUU –
3’). Nirenberg then added the synthesized mRNA to cell extracts that were capable
of translation.
a) What polypeptide(s) would you expect to find from the translation of this strand?
Explain.
4. In a similar experiment, Gobind Khorana synthesized an mRNA strand that was
composed of repeating units of “5’-UAC-3’.” He then translated the mRNA using a
technique similar to Nirenberg.
a) What polypeptide(s) would you expect to find from the translation of this strand?
(hint: assume the mRNA was synthesized in such a way that the first base could
not be established). Explain.
b) Would you expect to see mixtures of amino acids in any of the translated
peptides? Why or why not?
Khorana also attempted to translate mRNAs composed of four base repeats. One
mRNA he attempted to translate, 5’-GUAA-3’, resulted in the formation of only triand di-peptides.
c) Why didn’t any protein longer than a tripeptide result? Explain.
5. Which of the following amino acids would Ile-tRNA synthetase have the most
difficulty discriminating against? Why?
O
H 2N
OH
H 2N
O
Phenylalanine
OH
H 2N
O
Leucine
OH
H 2N
O
Valine
OH
H 2N
O
Asparagine
6. Below is the DNA sequence of a gene involved in leucine biosynthesis in E. coli
1
l
25
l
50
+1
l
5’-TAGTGTATTGACATGATAGAAGCACTCTACTATATTCTCAATAGGTCCACGGGTCCACCG
5’-ATCACATAACTGTACTATCTTCGTGAGATGATATAAGAGTTATCCAGGTGCCCAGGTGGC
100
l
75
l
AATATGACTCACATCGTTCGCTTTATCGGTCTACTACTACTAAACGCATCTTCTTTGCGCGGT
TTATACTGAGTGTAGCAAGCGAAATAGCCAGATGATGATGATTTGCGTAGAAGAAACGCGCCA
125
l
175
l
150
l
AGACGAGTGAGCGGCATCCAGCATTAACCCACAGCCGCCACTTCCGCTGGCGGCATTTTAAA-3’
TCTGCTCACTCGCCGTAGGTCGTAATTGGGTGTCGGCGGTCAAGGCGACCGCCGTAAAATTT-5’
a) Please write the first 18 bases of the mRNA that would be transcribed, 5’ to 3’.
b) What will be the first five amino acids of the protein derived from the mRNA
(please denote N and C ends of the peptide and give your answer in both the
three letter code and single letter code)
c) What will be the last five amino acids of the protein derived from the mRNA
(please denote N and C ends of the peptide and give your answer in both the
three letter code and single letter code)?
d) What is the length of this protein (in amino acids)?
e) For each of the following nucleotide changes, please determine their effects on
the codon, amino acid specified, and on the resulting protein (truncation, single
amino acid change, radically different protein, or no change).
Mutation
Example:
C75→deletion
T74→deletion
G76→A76
A124→T124
C81→T81
C100→nothing
G175→A175
Change in
Codon
ATC→ATG
Change in
Amino Acid
Ile →Met
Effect on Protein
Radically different
protein
1. HMG CoA Reductase
a) There are introns in the HMG CoA gene.
b) It would be much longer than it actually is.
8275 amino acids (1 remaining nucleotide).
c) 2664 nucleotides not including the stop codon. If they include the stop
codon (2667) it is fine. It is also okay if they add three for the start
codon (2670) and say this methionine is sometimes cleaved off.
d) Yes, there is a disparity. The start site of transcription is not the same
as the start site for translation. The termination of translation is also
not the same as the termination of transcription.
e) This tells you that the coding region of this gene in yeast does not have
any introns.
2. The polypeptide extends by adding amino acids onto the C-terminal, so
in the C-terminal direction.
3. a) The codon “UUU” encodes phenylalanine, so one would expect a
polypeptide composed of entirely of phenylalanine… this is exactly what
Nirenberg saw. Since all the bases are identical, “frames” are not of any
concern
4. a) You would expect to find poly(tyrosine), poly(threonine), and
poly(leucine), for their respective codons of UAC, ACU, and CUA.
b) No, you would not; since the templates repeat every 3-bases, there is
no change in codons within the reading frame. Thus, you would not
expect to see any heteropeptides formed
c) The GUAA repeat shifts translation between 4 different codons: GUA,
AGU, AAG, and UAA. GUA encodes for valine, AGU for serine, and AAG
for Lys. However, UAA is a stop codon; therefore, regardless of which
frame translation began, a stop codon was always reached within four
codons. Thus, the longest possible translated peptide was a tripeptide.
5. Valine and leucine since they are of a similar size and have similar
chemical properties (both have alkyl sidechains made of carbon and
hydrogen atoms).
6. Leucine biosynthesis in E. coli
a) 5’-AGGUCCACGGGUCCACCG-3’
b) N-Met-Thr-His-Ile-Val-C
N-MTHIV-C
c) N-Ser-Gly-Ile-Gln-His-C
N-SGIQH-C
d) 28
e) Below
Mutation
Example:
C75→deletion
T74→ deletion
Change in Codon
AUC→AUG
Change in Amino Acid
Ile→Met
AUC→ACG
Ile→Thr
G76→A76
GUU→AUU
Val→Ile
A124→T124
C81→T81
C100→ deletion
G175→A175
AGA→ UGA
CGC→ CGU
CTA→ UAA
No codon
Arg→ stop
Arg→ Arg, none
Leu→stop
None-not in coding
region
Effect on Protein
Radically different
protein
Radically different
protein
Single amino acid
change
truncation
No change
truncation
No change